FLEX Opentrons Flex Open Source Liquid Handling Robot

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Opentrons Flex

Specifications:

• General Specifications: Lorem ipsum dolor sit
  amet, consectetur adipiscing elit. Sed do eiusmod tempor incididunt
  ut labore et dolore magna aliqua.

• Environmental Specifications: Lorem ipsum
  dolor sit amet, consectetur adipiscing elit. Sed do eiusmod tempor
  incididunt ut labore et dolore magna aliqua.

• Certifications: Lorem ipsum dolor sit amet,
  consectetur adipiscing elit. Sed do eiusmod tempor incididunt ut
  labore et dolore magna aliqua.

• Serial Number: XXX-XXXX-XXXX

Product Usage Instructions:

1. Instrument Installation and Calibration:

Follow the steps outlined in the manual for pipette and gripper
installation.

2. Relocation:

For short moves, refer to section 2.5 in the manual. For
long-distance moves, follow the guidelines provided. General moving
advice is also available.

3. Connections:

Ensure proper power connection as detailed in the manual.
Connect USB and auxiliary devices as needed. Network connections
should be established following the instructions.

4. Protocol Designer:

Understand the requirements for Protocol Designer and learn how
to design new protocols or modify existing ones as per your lab
requirements.

5. Python Protocol API:

Explore writing and running scripts using Python Protocol API.
Discover Python-exclusive features for enhanced functionality.

6. OT-2 Protocols:

Learn about OT-2 Python protocols, OT-2 JSON protocols, and
Magnetic Module protocols for different types of experiments.

Frequently Asked Questions (FAQ):

Q: How do I troubleshoot if the robot is not moving as

expected?

A: Check the power connection, ensure proper calibration of
instruments, and verify that there are no obstructions in the
robot’s path.

Q: Can I use custom pipettes with Opentrons Flex?

A: Opentrons recommends using compatible pipettes for optimal
performance and accuracy.

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Opentrons Flex
Instruction Manual
Opentrons Labworks Inc.
December 2023

© OPENTRONS 2023 Opentrons FlexTM (Opentrons Labworks, Inc.) Registered names, trademarks, etc. used in this document, even when not specifically marked as such, are not to be considered unprotected by law.

Table of Contents
Preface . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .9 Structure of this manual . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9 Notes and warnings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10
Chapter 1: Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11 1.1 Welcome to Opentrons Flex . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11 What’s new in Flex. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11 Flex workstations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12 1.2 Safety information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16 Safety symbols. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16 Electrical safety warnings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17 Additional safety warnings . . . . . . . . . . . . . . . . . . . . . . . . . . . .18 Safety cautions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .18 Biological safety . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19 Toxic fumes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19 Flammable liquids . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19 1.3 Regulatory compliance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .20 Safety . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .20 Electromagnetic compatibility . . . . . . . . . . . . . . . . . . . . . . . . .20 FCC warnings and notes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .20 Canada ISED compliance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21 Environmental warning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21 Wi-Fi precertification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21
Chapter 2: Installation and Relocation . . . . . . . . . . . . . . . . . . . . . . . 22 2.1 Safety and operating requirements . . . . . . . . . . . . . . . . . . . . .22 Where to place Opentrons Flex . . . . . . . . . . . . . . . . . . . . . . . .22 Power consumption . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .24 Environmental conditions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .24 2.2 Unboxing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .25 Effort and time required . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .25 Crate and packing material . . . . . . . . . . . . . . . . . . . . . . . . . . . .25 Product elements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .26 Part 1: Remove the crate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .27 Part 2: Release the Flex. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .29 Part 3: Final assembly and power on . . . . . . . . . . . . . . . . . . .32

2.3 First run . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35 Power on . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35 Connect to a network or computer . . . . . . . . . . . . . . . . . . . . .36 Install software updates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .37 Attach Emergency Stop Pendant . . . . . . . . . . . . . . . . . . . . . . 38 Give your robot a name . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 38
2.4 Instrument installation and calibration . . . . . . . . . . . . . . . . . 38 Pipette installation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .39 Gripper installation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .40
2.5 Relocation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .40 Short moves. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 41 Long-distance moves . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 41 General moving advice . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .42 Final thoughts about moving. . . . . . . . . . . . . . . . . . . . . . . . . . .44
Chapter 3: System Description. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45 3.1 Physical components . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .45 Frame and enclosure . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .46 Deck and working area . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .46 Staging area . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .47 Deck fixtures. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 48 Waste chute. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .49 Staging area slots . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .49 Movement system . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .51 Touchscreen and LED displays . . . . . . . . . . . . . . . . . . . . . . . . .51 3.2 Pipettes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53 Pipette specifications. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 54 Pipette calibration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 55 Pipette tip rack adapter. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 55 Partial tip pickup . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .56 Pipette sensors. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .56 Pipette firmware updates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .57 3.3 Gripper . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .57 Gripper specifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58 Gripper calibration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58 Gripper firmware updates . . . . . . . . . . . . . . . . . . . . . . . . . . . . 58 3.4 Emergency Stop Pendant . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .59 When to use the E-stop . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .59 Engaging and releasing the E-stop . . . . . . . . . . . . . . . . . . . . .60

3.5 Connections . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61 Power connection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61 USB and auxiliary connections . . . . . . . . . . . . . . . . . . . . . . . . .62 Network connections . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .62
3.6 System specifications. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .63 General specifications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .63 Environmental specifications . . . . . . . . . . . . . . . . . . . . . . . . . .64 Certifications. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .64 Serial number . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .64
Chapter 4: Modules . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 66 4.1 Supported modules . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .66 4.2 Module caddy system . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .67 4.3 Module calibration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .68 When to calibrate modules. . . . . . . . . . . . . . . . . . . . . . . . . . . . .69 How to calibrate modules . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .69 4.4 Heater-Shaker Module GEN1 . . . . . . . . . . . . . . . . . . . . . . . . . .70 Heater-Shaker features. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .70 Heater-Shaker specifications . . . . . . . . . . . . . . . . . . . . . . . . . .72 4.5 Magnetic Block GEN1 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .73 Magnetic Block features. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .73 Magnetic Block specifications . . . . . . . . . . . . . . . . . . . . . . . . .74 4.6 Temperature Module GEN2 . . . . . . . . . . . . . . . . . . . . . . . . . . . .75 Temperature Module features . . . . . . . . . . . . . . . . . . . . . . . . .75 Temperature Module specifications . . . . . . . . . . . . . . . . . . . .77 4.7 Thermocycler Module GEN2. . . . . . . . . . . . . . . . . . . . . . . . . . . .77 Thermocycler features . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .78 Thermocycler specifications . . . . . . . . . . . . . . . . . . . . . . . . . . .79
Chapter 5: Labware . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 80 5.1 Labware concepts. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .80 Labware as hardware . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .80 Labware as data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .80 Custom labware . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .81 5.2 Reservoirs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .82 Single-well reservoirs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .82 Multi-well reservoirs. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .82 Reservoirs and API definitions . . . . . . . . . . . . . . . . . . . . . . . . 83 Custom reservoir labware. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 83 5.3 Well plates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 83

6-well plates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84 12-well plates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84 24-well plates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84 48-well plates. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 84 96-well plates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 85 384-well plates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .86 Well plate adapters. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .86 Well plates and API definitions. . . . . . . . . . . . . . . . . . . . . . . . .87 Custom well plate labware. . . . . . . . . . . . . . . . . . . . . . . . . . . . .87 5.4 Tips and tip racks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 88 Tip racks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 88 Tip­pipette compatibility . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89 Tip rack adapter . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 89 5.5 Tubes and tube racks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .90 Tube and rack combinations . . . . . . . . . . . . . . . . . . . . . . . . . . . 91 6-tube racks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 91 10-tube racks. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 91 15-tube racks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .92 24-tube racks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .92 Tube rack API definitions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .92 Custom tube rack labware . . . . . . . . . . . . . . . . . . . . . . . . . . . . .93 5.6 Aluminum blocks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .93 Flat bottom plate . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .93 24-well aluminum block . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .94 96-well aluminum block . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .94 Standalone adapters . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .94 Aluminum block labware combinations . . . . . . . . . . . . . . . .94 24-well aluminum block labware combinations . . . . . . . . .95 96-well aluminum block labware combinations . . . . . . . . .95 5.7 Labware and the Opentrons Flex Gripper . . . . . . . . . . . . . . .96 5.8 Custom labware definitions . . . . . . . . . . . . . . . . . . . . . . . . . . . .96 Creating custom labware definitions . . . . . . . . . . . . . . . . . . .97 JSON labware schema . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .99 JSON labware definitions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 104 Chapter 6: Protocol Development . . . . . . . . . . . . . . . . . . . . . . . . . . .105 6.1 Pre-made protocols . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 105 Protocol Library . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 105 Custom Protocol Development service . . . . . . . . . . . . . . . . 107

6.2 Protocol Designer. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 109 Protocol Designer requirements . . . . . . . . . . . . . . . . . . . . . . 109 Designing a protocol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 110 Modifying existing protocols . . . . . . . . . . . . . . . . . . . . . . . . . . 114
6.3 Python Protocol API . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 115 Writing and running scripts . . . . . . . . . . . . . . . . . . . . . . . . . . . 115 Python-exclusive features . . . . . . . . . . . . . . . . . . . . . . . . . . . . 116
6.4 OT-2 protocols. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 117 OT-2 Python protocols. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 118 OT-2 JSON protocols . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 120 Magnetic Module protocols . . . . . . . . . . . . . . . . . . . . . . . . . . . 120
Chapter 7: Software and Operation. . . . . . . . . . . . . . . . . . . . . . . . . . 121 7.1 Touchscreen operation. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 121 Robot dashboard . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 121 Protocol management . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 122 Protocol details . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 124 Run setup . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 127 Labware Position Check . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 128 Run progress. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 130 Run completion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 131 Instrument management. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 132 Robot settings. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 133 Deck configuration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 136 7.2 Opentrons App. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 139 App installation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 139 Transferring protocols to Flex. . . . . . . . . . . . . . . . . . . . . . . . . 140 Module status and controls . . . . . . . . . . . . . . . . . . . . . . . . . . . 142 Recent protocol runs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 143 7.3 Advanced operation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 144 Jupyter Notebook . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 144 Command-line operation over SSH . . . . . . . . . . . . . . . . . . . . 145
Chapter 8: Maintenance and Service . . . . . . . . . . . . . . . . . . . . . . . .147 8.1 Cleaning your Flex . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 147 Before you begin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 147 What you can clean . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 147 Cleaning solutions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 148 Frame and window panel cleaning . . . . . . . . . . . . . . . . . . . . 148 Deck cleaning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 148

Gantry cleaning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 149 Waste chute cleaning. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 149 8.2 Cleaning pipettes and tips . . . . . . . . . . . . . . . . . . . . . . . . . . . . 149 Pipette decontamination . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 150 Cleaning pipette tips . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 151 8.3 Cleaning the gripper . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 152 8.4 Cleaning modules. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 153 General module cleaning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 153 Thermocycler seals. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 154 8.5 Autoclave-safe labware . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 154 8.6 Servicing Flex . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 155 Opentrons services . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 155 Installation qualification and operation qualification . . . 155 Preventative maintenance . . . . . . . . . . . . . . . . . . . . . . . . . . . . 156 Warranty . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 157 Appendix A: Glossary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .158 Appendix B: Additional Documentation . . . . . . . . . . . . . . . . . . . . . .170 B.1 Opentrons Knowledge Hub . . . . . . . . . . . . . . . . . . . . . . . . . . . . 170 B.2 Python Protocol API documentation . . . . . . . . . . . . . . . . . . . 170 B.3 Opentrons HTTP API reference. . . . . . . . . . . . . . . . . . . . . . . . 171 B.4 Developer documentation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 171 Appendix C: Open-Source Software . . . . . . . . . . . . . . . . . . . . . . . . .172 C.1 Opentrons on GitHub . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 172 C.2 Opentrons monorepo . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 173 C.3 Other repositories . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 175 Appendix D: Support and Contact Information . . . . . . . . . . . . . . .176 D.1 Sales . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 176 D.2 Support . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 176 D.3 Business information . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 177

Preface
Welcome to the instruction manual for the Opentrons Flex liquid handling robot. This manual guides you through just about everything you need to know to set up and use Flex, focusing on topics that are most relevant to everyday users of Flex in a lab environment.
Structure of this manual
Opentrons Flex is a complex system, so there are many different paths to learning everything it can do. Feel free to jump straight to the chapter that addresses whatever topic you’re curious about! For example, if you already have a Flex set up in your lab, you can skip past the Installation and Relocation chapter.
If you prefer a guided approach, this manual is structured so you can follow it from beginning to end.
Learn about Flex. The distinguishing features of Flex are listed in Chapter 1: Introduction. The introduction also includes important safety and regulatory information.
Get started with Flex. If you need to set up your Flex, follow the detailed instructions in Chapter 2: Installation and Relocation. Then familiarize yourself with the components of Flex in Chapter 3: System Description.
Set up your deck. Configuring the deck enables different scientific applications on Flex. Chapter 4: Modules describes Opentrons peripherals that you can install into or on top of the deck to perform specific scientific tasks. Chapter 5: Labware explains how to work with equipment for holding liquids.
Run a protocol. The core use of Flex is running standardized scientific procedures, known as protocols. Chapter 6: Protocol Development offers several ways to get ready-made protocols or design them yourself. To run your protocol, follow the instructions in Chapter 7: Software and Operation, which also has instructions for performing other tasks and customizing your robot’s settings.
Keep Flex running. Follow the advice in Chapter 8: Maintenance and Service to keep your Flex clean and running optimally. Or sign up for one of the Opentrons services listed there and let us take care of Flex for you.
Learn even more. Still need something else? Consult the appendices. Appendix A: Glossary defines Flex-related terms. Appendix B: Additional Documentation points you to even more resources for Opentrons products and writing code to control Flex.

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Appendix C: Open-Source Software explains how Opentrons software is hosted on GitHub as a resource for both developers and non-developers.
Appendix D: Support and Contact Information lists how to get in touch with Opentrons if you need assistance beyond what our documentation provides.
Notes and warnings
Throughout this manual, you’ll find specially formatted note and warning blocks. Notes provide helpful information that may not be obvious in the ordinary course of using Flex. Pay special attention to warnings–they are only used in situations where you run the risk of personal injury, damage to equipment, loss or spoilage of samples or reagents, data loss, or other harm. Notes and warnings look like this:
Sample Note: This is something you ought to know, but it doesn’t pose any danger.
Sample Warning: This is something you need to know because there is risk associated with it.

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Introduction

This chapter introduces you to the Opentrons Flex ecosystem, including the overall system design and available workstation configurations. It also includes important compliance and safety information, which you should review before setting up your Opentrons Flex robot. For more details on the features of Opentrons Flex, see the System Description chapter.

1.1 Welcome to Opentrons Flex
Opentrons Flex is a liquid-handling robot designed for high throughput and complex workflows. The Flex robot is the base of a modular system that includes pipettes, a labware gripper, deck fixtures, on-deck modules, and labware — all of which you can swap out yourself. Flex is designed with a touchscreen so you can work with it directly at the lab bench, or you can control it from across your lab with the Opentrons App or our open-source APIs.
Flex workstations come with all of the equipment — robot, hardware, and labware — that you need to get started automating common lab tasks. For other applications, Opentrons Flex runs on fully open-source software and firmware, and is reagent- and labware-agnostic, giving you control over how you design and run your protocols.

What’s new in Flex
Opentrons Flex is part of the Opentrons liquid handler series of robots. Users of Opentrons Flex may be familiar with the Opentrons OT-2, our personal pipetting robot. Flex goes beyond the capabilities of OT-2 in several key areas, delivering higher throughput and walkaway time.

Feature Pipette throughput
Pipette and tip capacities

Description
Flex pipettes have 1, 8, or 96 channels. The 96-channel pipette operates on 12 times as many wells at once as the largest OT-2 pipette.
Flex pipettes have larger volume ranges (1­50 µL, 5­1000 µL) and can all work with any volume of Opentrons Flex tips. This is an improvement over OT-2 pipettes, which have smaller ranges and must use tips with a matching volume range.

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Gripper Automated calibration Touchscreen Module caddies Deck slot coordinates Movable trash Size and weight

The Opentrons Flex Gripper picks up and moves labware around the deck automatically, without user intervention. The gripper enables more complex workflows within a single protocol run.
Positional calibration of Flex pipettes and the gripper is fully automated. Press one button, and the instrument will move to precision-machined points on the deck to determine its exact position, saving that data for use in your protocols.
Flex has its own touchscreen interface that lets you control it directly, in addition to using the Opentrons App. Use the touchscreen to start protocol runs, check job status, and change settings right on the robot.
Flex modules fit into caddies that occupy space below the deck. Caddies place your labware closer to the deck surface and allow for below-deck cable routing. Caddies enable even more module and labware configurations on the deck.
Deck slots on Flex are numbered with a coordinate system (A1­D4) which is similar to how wells are numbered on labware.
The trash bin can go in multiple deck locations on Flex. The default location (slot A3) is the recommended position. You can also use the gripper to dispose of trash in the optional waste chute.
Flex is a bit bigger and much heavier than OT-2. Installation tasks on Flex require the assistance of a lab partner.

A detailed comparison of robot technical specifications is available on the Opentrons website.
Both Flex and OT-2 robots run on our open-source software, and the Opentrons App can control both types of robots at once. While OT-2 protocols can’t be run directly on Flex, it’s straightforward to adapt them (see the OT-2 Protocols section of the Protocol Development chapter for details).

Flex workstations
Opentrons Flex workstations include the Flex robot, accessories, pipettes and gripper, on-deck modules, and labware needed to automate a particular application. All workstation components are modular. If you need to change applications, you can add or swap in other Flex hardware and compatible consumables.

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NGS WORKSTATION
The Opentrons Flex NGS Workstation automates NGS library prep. It can automate pre-sequencing workflows using any leading reagent system, including fragmentation- and tagmentation-based library prep.
In addition to the Flex robot, the NGS Workstation includes:
Gripper Choice of pipette configuration
Two 8-Channel Pipettes (1­50 µL and 5­1000 µL) 96-Channel Pipette (5­1000 µL) Waste Chute Magnetic Block Temperature Module Thermocycler Module Labware kit with filter tips, microcentrifuge tubes, reservoirs, and PCR plates
PCR WORKSTATION
The Opentrons Flex PCR Workstation automates PCR setup and thermocycling workflows for up to 96 samples. It can aliquot chilled reagents and samples into a 96-well PCR plate. With the addition of the automated Thermocycler Module, use the gripper to load the plate into the Thermocycler, and then run your chosen PCR program.
In addition to the Flex robot, the PCR Workstation includes:
Gripper Choice of pipette configuration
1-Channel Pipette (1­50 µL) and 8-Channel Pipette (1­50 µL) 96-Channel Pipette (5­1000 µL) Waste Chute Temperature Module Labware kit with filter tips, microcentrifuge tubes, reservoirs, and PCR plates

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NUCLEIC ACID EXTRACTION WORKSTATION
The Opentrons Flex Nucleic Acid Extraction Workstation automates DNA/RNA isolation and purification. It uses the Magnetic Block for separation of magnetic beads, and the Heater-Shaker for sample lysis and resuspension of magnetic beads.
In addition to the Flex robot, the Nucleic Acid Extraction Workstation includes:
Gripper Choice of pipette configuration
1-Channel Pipette (5­1000 µL) and 8-Channel Pipette (5­1000 µL) 96-Channel Pipette (5­1000 µL) Waste Chute Magnetic Block Heater-Shaker Module Labware kit with filter tips, reservoirs, PCR plates, and deep well plates
MAGNETIC BEAD PROTEIN PURIFICATION WORKSTATION
The Opentrons Flex Magnetic Bead Protein Purification Workstation automates small-scale protein purification and proteomics sample prep for up to 96 samples. It is compatible with many popular magnetic-bead-based reagents.
In addition to the Flex robot, the Protein Purification Workstation includes:
Gripper Choice of pipette configuration
1-Channel Pipette (5­1000 µL) and 8-Channel Pipette (5­1000 µL) 96-Channel Pipette (5­1000 µL) Waste Chute Magnetic Block Heater-Shaker Module Labware kit with filter tips, reservoirs, PCR plates, and deep well plates
FLEX PREP WORKSTATION
The Opentrons Flex Prep Workstation automates simple pipetting workflows. Configure the workstation with 1-Channel and 8-Channel Pipettes to perform tasks such as sample transfer, sample duplication, and

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reagent aliquoting. Configure the workstation with the 96-Channel Pipette to perform high-throughput reagent aliquoting and plate stamping.
In addition to the Flex robot, the Flex Prep Workstation includes:
Choice of pipette configuration: 1-Channel Pipette (5­1000 µL) and 8-Channel Pipette (5­1000 µL) 96-Channel Pipette (5­1000 µL)
Labware kit with filter tips, microcentrifuge tubes, and reservoirs
PLASMID PREP WORKSTATION
The Opentrons Flex Plasmid Prep Workstation automates magnetic-bead-based plasmid extraction and purification workflows. This workstation is equipped with high-volume pipettes, a Heater-Shaker Module, and a Magnetic Block to accommodate most bead-based chemistry.
In addition to the Flex robot, the Plasmid Prep Workstation includes:
Gripper 1-Channel Pipette (5­1000 µL) and 8-Channel Pipette (5­1000 µL) Waste Chute Magnetic Block Heater-Shaker Module Labware kit with filter tips, microcentrifuge tubes, reservoirs, PCR plates, and deep well plates
SYNBIO WORKSTATION
The Opentrons Flex SynBio Workstation automates a variety of synthetic biology workflows such as DNA synthesis and cloning. It uses the Magnetic Block and Temperature Module to support most bead-based chemistry. Add the Thermocycler Module to perform heated lid incubations and amplifications.
In addition to the Flex robot, the SynBio Workstation includes:
Gripper 1-Channel Pipette (5­1000 µL) and 8-Channel Pipette (5­1000 µL) Magnetic Block Temperature Module

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Labware kit with regular and filter tips, microcentrifuge tubes, reservoirs, PCR plates, and deep well plates

1.2 Safety information
The Opentrons Flex liquid handling robot has been designed for safe operation. Refer to the specifications and compliance guidelines in this section to ensure safe usage of your Flex. These guidelines cover safe use of input and output connections for the product, including the power and data connections, as well as warning labels found on the Flex robot and related hardware. Using the device in a manner other than those specified in this manual may put the user and equipment at risk.

Safety symbols
Various labels on the Flex and in this manual warn you about sources of potential injury or harm.

Symbol

Description
Warning: Alerts users to Potentially hazardous conditions. Actions that may result in personal injury or death.
Caution: Cautions users against Damage to the equipment. Lost or corrupted data. Unrecoverable interruption of the operation being performed.
Electrical shock: Identifies instrument components that might pose a risk of electrical shock if the instrument is handled improperly.

Hot surface: Identifies instrument components that pose a risk of personal injury due to high heat/temperatures if the instrument is handled improperly.

Pinch point: Identifies instrument components that can pose a risk of personal injury when in motion.

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You will find the following labels on the Flex:
Intellectual property labels Regulatory compliance labels (e.g., ETL) Electrical hazard labels General warning labels Product labels Pinch point labels High voltage labels Power rating labels

Electrical safety warnings
Always observe the following electrical safety warnings:

Symbol

Description
Plug the robot into a grounded, Class 1 circuit. See the Power Connection section in the System Description chapter.

Do not connect (plug in), disconnect (unplug), or use AC power cables if: The cable is frayed or damaged. Other attached cables, cords, or receptacles are frayed or damaged.
Using damaged power cords can cause an electric shock hazard resulting in serious injury or damage to the robot.
Do not replace the AC power cable unless at the direction of Opentrons Support.

For more information on electrical requirements, see the Power Consumption section of the Installation and Relocation chapter.

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Additional safety warnings
Always observe the following additional safety warnings:

Symbol

Description
Opentrons Flex has not been certified for use with explosive or flammable liquids. Do not load plates, tubes, or vials containing explosive or flammable liquids into the robot or otherwise operate the instrument with explosive or flammable liquids in the enclosure.
Use good laboratory practices and follow the manufacturer’s precautions when working with chemicals. Opentrons is not responsible or liable for any damages because of, or as a result of, the use of hazardous chemicals.

The Flex weighs 88.5 kg (195 lbs). As a result, it requires two people to lift and move it safely. See the Relocation section in the Installation and Relocation chapter.

The Flex should be placed on a surface capable of supporting its weight of 88.5 kg (195 lbs) with sufficient surface area to accommodate the robot plus its minimum clearance distance (20 cm/8 in). See the Safety and Operating Requirements section in the Installation and Relocation chapter.
The Flex can emit vibrations while in operation. Place the robot on a surface that is sturdy, level, and water-resistant with cross-bracing or welded joints. See the Safety and Operating Requirements section in the Installation and Relocation chapter.

Safety cautions
To help protect the Flex from damage, follow these precautions:

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Symbol

Description
Use labware that is ANSI/SLAS-compliant or approved by Opentrons. See the Labware chapter.
Keep corrosive materials, agents, or otherwise damaging materials away from the robot.

Biological safety
Treat specimens and reagents containing materials taken from humans as potentially infectious agents. Opentrons recommends using safe laboratory procedures as explained in Biosafety in Microbiological and Biomedical Laboratories (BMBL) 6th Edition.
Under normal circumstances, the Flex does not create detectable aerosols from source liquids. However, under certain conditions, it is possible to generate aerosols from source liquids. When operating with biosafety level 2 or greater source liquids, consider taking precautions against aerosol exposure, in accordance with your local regulatory bodies. To minimize the potential risk of aerosol exposure from the robot, ensure that you:
Perform maintenance as described in the Maintenance and Service chapter. Properly install and secure all instrument covers, pipettes, modules, and labware. Use proper pipetting technique to aid in the mitigation of aerosols.
Toxic fumes
If you’re working with volatile solvents or toxic substances, use an efficient laboratory ventilation system to remove any vapors that may be produced.
Flammable liquids
The Flex has not been evaluated for use with flammable liquids and should not be used with flammable liquids.

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1.3 Regulatory compliance
Opentrons Flex complies with all applicable requirements of the following safety and electromagnetic standards.

Safety
Rule ID IEC/UL/CSA 61010-1 IEC/UL/CSA 61010-2-051

Title
Safety Requirements for Electrical Equipment for Measurement, Control, and Laboratory Use ­Part 1: General Requirements
Particular requirements for laboratory equipment for mixing and stirring

Electromagnetic compatibility

Rule ID EN/BSI 61326-1
FCC 47 CFR Part 15 Subpart B Class A IC ICES-003

Title
Electrical Equipment for Measurement, Control, and Laboratory Use ­ EMC Requirements ­ Part 1: General Requirements Unintentional Radiators
Spectrum Management and Telecommunications ­InterferenceCausing Equipment Standard ­ Information Technology Equipment (Including Digital Apparatus)

FCC warnings and notes
Warning: Changes or modifications to this unit not expressly approved by Opentrons could void the user’s authority to operate the equipment.
This device complies with Part 15 of the FCC rules. Operation is subject to the following:

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This device may not cause harmful interference. This device must accept any interference received, including interference that may cause undesired
operation.
Note: This equipment has been tested and found to comply with the limits for a Class A digital device, pursuant to Part 15 of the FCC rules. These limits are designed to provide reasonable protection against harmful interference when the equipment is operated in a commercial environment. This equipment generates, uses, and can radiate radio frequency energy and, if not installed and used in accordance with the instruction manual, may cause harmful interference to radio communications. Operation of this equipment in a residential area is likely to cause harmful interference in which case the user will be required to correct the interference at their own expense.
Canada ISED compliance
Canada ICES-003(A) / NMB-003(A)
This product meets the applicable Innovation, Science and Economic Development Canada technical specifications.
Le présent produit est conforme aux spécifications techniques applicables d’Innovation, Sciences et Développement économique Canada.
Environmental warning
Warning: Cancer and Reproductive Harm ­ www.P65Warnings.ca.gov
Wi-Fi precertification
The Wi-Fi module is precertified for use in many regions:
United States (FCC): FCC Identifier UAY-W8997-M1216 European Economic Area (CE): No public identifier (self-declaration) Canada (IC): Hardware Version Identification Number W8997-M1216 Japan (TELEC): Certified number 020-170034 India (WPC): Registration number ETA-SD-20191005525 (self-declaration)

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CHAPTER 2
Installation and Relocation
This chapter describes how to prepare your lab for Opentrons Flex, how to set up the robot, and how to move it if necessary. Before taking delivery of your Flex, make sure that your lab or facility meets all the criteria in the Safety and Operating Requirements section. When it’s time to get your Flex up and running, follow the detailed instructions in the Unboxing, First Run, and Instrument Installation and Calibration sections, or use the Opentrons Onsite Support Set Up service. And if you ever need to move your Flex to a new location, near or far, follow the steps in the Relocation section.
2.1 Safety and operating requirements
Where to place Opentrons Flex
Space is a valuable commodity in almost every lab. Your Flex is going to need some–but not too much, as it’s designed to fit on half of a standard lab bench. Make sure that you have a space that meets the following criteria.
Bench surface: Stationary, sturdy, level, water-resistant surface. Tables or benches with wheels (even locking wheels) are not recommended. Flex moves quickly and has a lot of mass, which can shake or imbalance lightweight or movable tables.
Weight bearing: The robot alone weighs 88.5 kg (195 lb) and should only be lifted by two people working together. Place the robot on a surface that can readily support its weight plus the weight of any modules, labware, liquids, or other lab equipment to be used in your applications.
Operating space: The robot’s base dimensions are 87 cm W x 69 cm D x 84 cm H (about 34″ x 27″ x 33″). Flex needs 20 cm (8″) of side and back clearance for cables, USB connections, and to dissipate exhaust from modules that heat and cool.
Warning: Do not position the sides or back of the Flex flush against a wall.

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84 cm 33″

87 cm 34″
Opentrons Flex base dimensions.

69 cm 27″

20 cm 8″

20 cm 8″

20 cm 8″

Top view of Opentrons Flex, showing minimum side and back clearance.

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Power consumption
Opentrons Flex should be connected to a wall outlet at or near the bench location where you install it. Only connect Flex to circuits that can accommodate its maximum power draw:
Input power: 36 VDC, 6.1 A Idle consumption: 30­40 W Typical consumption: 50­100 W (during protocol run) Maximum consumption: Approximately 120 W
Exact power consumption depends on:
The amount and type of movement executed during a protocol. The amount of time the robot spends idle. The status of the lights on the robot. How many instruments are attached.
Remember to account for other electronics that consume power on the same circuit, including Flex modules with their own power supplies. For example, the Thermocycler Module has a maximum power consumption (630 W) that is much greater than the Flex robot itself. If necessary, consult the manager of your facility to make sure it meets your equipment’s power requirements.

Environmental conditions
Environmental conditions for recommended use, acceptable use, and storage vary:

Recommended for system operation

Acceptable for system operation

Ambient temperature +20 to +25 °C

+2 to +40 °C

Relative humidity Altitude

40­60%, non-condensing
Approximately 500 m above sea level

30­80%, non-condensing (below 30 °C)
Up to 2000 m above sea level

Storage and transportation
-10 to +60 °C
10­85%, non-condensing (below 30 °C)
Up to 2000 m above sea level

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Opentrons has validated the performance of Opentrons Flex in the conditions recommended for system operation, and operation in those conditions should provide optimal results. Flex is safe to use in conditions acceptable for system operation, but results may vary. Do not power on or use Flex in conditions outside of those bounds. The storage and transportation conditions only apply when the robot is completely disconnected from power and other equipment.
2.2 Unboxing
Congratulations! Your Opentrons Flex has arrived and you’ve prepared a space for it in your lab. Let’s open that monster crate, remove the robot, and prepare it for operation. The information in this section provides a parts list and instructions that walk you through the steps required to get the Flex unboxed, set up, and ready for use. We’ve divided the setup procedure into three parts:
Part 1 covers disassembling the crate. Part 2 covers detaching the Flex from the crate and moving it to a final assembly location. Part 3 covers final assembly and powering on the robot for the first time.
Effort and time required
You’ll want to ask a lab partner to assist with the unboxing, lifting, moving, and assembly process. You’ll need to budget about 30 minutes to an hour for this effort.
Note: The Flex requires two people to lift it properly. Also, lifting and carrying the Flex by its handles is the best way to move the robot.
Crate and packing material
Unpacking a Flex gives you an awesome robot, but you’re also left with several large crate panels along with assorted shipping components and padding. While you could discard this material, we encourage you to keep these items if storage space is available. The packaging is reusable, which helps prepare your Flex for shipping if you ever need to send it somewhere else (e.g., to a conference or a new facility) in the future.

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Product elements
The Flex ships with the components listed below. Pipettes, the gripper, and modules come in separate packaging from the main Flex crate, even if you purchased them together as a workstation.

(1) Opentrons Flex robot

(1) USB cable

(1) Power cable

(1) Ethernet cable

(5) L-keys (12 mm hex, 1.5 mm hex, 2.5 mm hex, 3 mm hex,
T10 Torx)

(1) Emergency Stop Pendant

(1) Deck slot with labware clips

(4) Spare labware clips

(1) Pipette calibration probe

(4) Carrying handles and caps

(1) Top window panel

(4) Side window panels

(1) 2.5 mm hex screwdriver

(1) 19 mm wrench

(16 + spares) Window screws (M4x8 mm flat head)
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(10) Spare deck slot screws (M4x10 mm socket head)

(12) Spare deck clip screws (M3x6 mm socket head)

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Part 1: Remove the crate
Opentrons ships your Flex in a sturdy plywood crate. The shipping crate uses hook and latch clamps to secure the top, side, and bottom panels together. Using latches, instead of nails or screws, means you won’t need a crowbar (or a lot of force) to disassemble the crate, and you can reassemble it later, if needed.
Note: Crate edges can get roughed up during shipping. You may want to use work gloves to protect your hands from wood splinters.
To release the latches, flip the latch tab up and turn it to the left (counterclockwise). This action moves the clamp arm out of its corresponding retaining bracket. You can then flip the latch arm away from the crate.
1 Unlock the eight latches holding the top to the sides.

2 Remove the top panel after releasing the latches.

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CHAPTER 2: INSTALLATION AND RELOCATION 3 Cut open the blue shipping bag, remove these items from the padding, and set them aside:
User Kit Power, Ethernet, and USB cables Emergency Stop Pendant
4 Remove the top piece of foam padding to expose the window panels. The padding protects the side and top panels.
5 Remove the window panels and set them aside. You’ll attach these later.

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6 Unlock the remaining 16 latches holding the side panels to each other and the base of the crate. 7 Remove the side panels and set them aside.
Part 2: Release the Flex
After completing the steps in Part 1, you should now see a robot that’s in a protective bag and attached to orange steel mounting components. The bag encloses the robot and protects it from the outside environment. Steel brackets secure the robot to the bottom of the crate. Two shipping frames support the robot, distributing its weight evenly, and keeping it rigid so it doesn’t warp during shipping. Continue to unpack the Flex and get it off the crate base.
8 Using the 19 mm wrench from the User Kit, unbolt the brackets from the crate bottom. You can discard the brackets, or save them for future use.

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CHAPTER 2: INSTALLATION AND RELOCATION 9 Pull or roll the shipping bag all the way down to expose the entire robot.
10 With help from your lab partner, grab the handholds in the orange shipping frames on either side of the robot’s base, lift the Flex off the crate base, and set it down on the floor. Save or discard the crate base and shipping frame.

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CHAPTER 2: INSTALLATION AND RELOCATION 11 Using the 12 mm hex L-key from the User Kit, remove the four bolts holding the shipping frames to
the Flex. Save or discard the frames and bolts.
12 Remove the four aluminum handles from the User Kit. Screw the handles into the same locations that held the 12 mm shipping frame bolts.

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13 With help from your lab partner, lift the Flex by its carrying handles and move it to a workbench for final assembly.

Part 3: Final assembly and power on
After moving the Flex to a temporary work area, or its permanent home, it’s time to put the finishing touches on your new robot.
14 If you have moved the robot to its final, working location, remove the carrying handles and replace them with the finishing caps. The caps close the handle openings in the frame and give the robot a clean appearance. Return the handles to the User Kit for storage.

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15 Retrieve the top and side panels from the packing foam you set aside after removing the crate top.
16 Fit the window panels to the Flex by following the labeling information on the front protective film. Then remove the protective film.
17 Using the beveled window screws and the 2.5 mm screwdriver from the User Kit, attach the window panels to the Flex. Make sure the beveled (V-shaped) holes in the window panels are facing out (towards you). This allows the screws to fit flush with the surface of the window.

Warning: Incorrectly orienting the panels can lead to damage. Excessive screw torque can crack the panels. Hand tighten the screws until the window panels are reasonably secure. This is not a trial of strength.

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CHAPTER 2: INSTALLATION AND RELOCATION 18 Using the 2.5 mm screwdriver from the User Kit, remove the locking screws from the gantry. These
screws prevent the gantry from moving while in transit. The gantry locking screws are located: On the left side rail near the front of the robot. Underneath the vertical gantry arm. On the right side rail near the front of the robot in an orange bracket. There are two screws here.
The gantry moves easily by hand after removing all the shipping screws. 19 Cut and remove the two rubber bands that hold the trash bin in place during shipping.

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20 Attach the power cord to Flex and plug it into the wall outlet. Make sure the deck area is free of obstructions. Flip the power switch on the back left of the robot. Once powered on, the gantry moves to its home location and the touchscreen displays additional configuration instructions.

Now that your Flex is out of the box and ready to go, continue to the First Run section below.
2.3 First run
Perform basic setup on the touchscreen before connecting any other hardware to your Flex. The robot will guide you through connecting to your lab network, updating to the latest software, and personalizing Flex by giving it a name.
Power on
When you power on Flex, the Opentrons logo will appear on the touchscreen. After a few moments, it will show the “Welcome to your Opentrons Flex” screen.
The Opentrons Flex welcome screen. You should only see this screen when you start your Flex for the first time.

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Connect to a network or computer
Follow the prompts on the touchscreen to get your robot connected so it can check for software updates and receive protocol files. There are three connection methods: Wi-Fi, Ethernet, and USB.

Network connection options. You need to have internet connectivity to set up Flex. Wi-Fi: Use the touchscreen to connect to Wi-Fi networks that are secured with WPA2 Personal authentication (most networks that only require a password to join fall under this category).
Note: Flex does not support captive portals (networks that don’t have a password but load a webpage to authenticate users after connecting).
You can also connect to an open Wi-Fi network, but this is not recommended.
Warning: Connecting to an open Wi-Fi network will allow anyone in range of the network signal to control your Opentrons Flex robot without authentication.

If you need to connect to a Wi-Fi network that uses enterprise authentication (including “eduroam” and similar academic networks that require a username and password), first connect to the Opentrons App by Ethernet or USB to complete initial setup. Then connect to the enterprise Wi-Fi network in the networking settings for your Flex. To access the networking settings:

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1. Click Devices in the left sidebar of the Opentrons App. 2. Click the three-dot menu () for your Flex and choose Robot Settings. 3. Click the Networking tab.
Select your network from the dropdown menu or choose “Join other network…” and enter its SSID. Choose the enterprise authentication method that your network uses. The supported methods are:
EAP-TTLS with TLS EAP-TTLS with MS-CHAP v2 EAP-TTLS with MD5 EAP-PEAP with MS- CHAP v2 EAP-TLS
Each of these methods requires a username and password, and depending on your exact network configuration may require certificate files or other options. Consult your facility’s IT documentation or contact your IT manager for details of your network setup.
Ethernet: Connect your robot to a network switch or hub with an Ethernet cable. You can also connect directly to the Ethernet port on your computer, starting in robot system version 7.1.0.
USB: Connect the provided USB A-to-B cable to the robot’s USB-B port and an open port on your computer. Use a USB B-to-C cable or a USB A-to-C adapter if your computer does not have a USB-A port.
To proceed with setup, the connected computer must have the Opentrons App installed and running. For details on installing the Opentrons App, see the App Installation section of the Software and Operation chapter.
Install software updates
Now that you’ve connected to a network or computer, the robot can check for software and firmware updates and download them if needed. If there is an update, it may take a few minutes to install. Once the update is complete, the robot will restart.

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Attach Emergency Stop Pendant
Connect the included Emergency Stop Pendant (E-stop) to an auxiliary port (AUX-1 or AUX-2) on the back of the robot.

Before and after connecting the Emergency Stop Pendant.
Attaching and enabling the E-stop is mandatory for attaching instruments and running protocols on Flex. For more information on using the E-stop during robot operation, see the Emergency Stop Pendant section of the System Description chapter.
Give your robot a name
Naming your robot lets you easily identify it in your lab environment. If you have multiple Opentrons robots on your network, make sure to give them unique names. Once you’ve confirmed your robot’s name, you’ll be taken to your Opentrons Flex Dashboard. Likely the next step you’ll want to take is attaching instruments, which is covered in the next section.
2.4 Instrument installation and calibration
After initial robot setup, the next step is to attach instruments to the robot and calibrate them.
To install an instrument, first tap on Instruments on the touchscreen or go to the Pipettes and Modules section of the device detail screen in the Opentrons App. Choose an empty mount and select either Attach Pipette or Attach Gripper. If the mount you want to use is already occupied, you need to detach the pipette or gripper first.

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Note: The overall installation process is the same regardless of whether you use the touchscreen or the Opentrons App. Whatever device you begin on will control the installation process until you complete or cancel it.
If you begin on the touchscreen, the app will show the robot as being “busy”. If you begin in the app, the touchscreen will show a modal indicating that instrument installation is in progress.
The exact installation process varies depending on the instrument you are attaching, as covered in the sections below. All instruments have an automated calibration procedure, which you should perform immediately after installation.
Pipette installation
When you install a pipette, you will be guided through the following steps on the touchscreen or in the Opentrons App.
1. CHOOSE PIPETTE TYPE Choose between 1- or 8-Channel Pipette and 96-Channel Pipette. Attaching the 96-Channel Pipette requires a few additional steps because it attaches to a special mounting plate that spans both pipette mounts.
2. PREPARE FOR INSTALLATION Remove labware from the deck and clean up the working area to make attachment and calibration easier. Also gather the needed equipment, such as the calibration probe, hex screwdriver, and mounting plate (for the 96-Channel Pipette).
3. CONNECT AND SECURE THE PIPETTE The gantry will move to the front of the robot so you can attach the pipette.
1- and 8-Channel Pipettes connect directly to a pipette mount. The 96-Channel Pipette requires a mounting plate. In order to attach the mounting plate, you must first disconnect the z-axis carriage for the right pipette mount.

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Connect the pipette to the chosen pipette mount and secure its screws.
4. RUN AUTOMATED CALIBRATION To calibrate the pipette, attach the calibration probe to the appropriate pipette nozzle. The pipette will automatically move to touch certain points on the deck and save these calibration values for future use. Once calibration is complete and you’ve removed the probe, the pipette will be ready for use in protocols.
Gripper installation
When you install the gripper, you will be guided through the following steps on the touchscreen or in the Opentrons App.
1. PREPARE FOR INSTALLATION Remove labware from the deck and clean up the working area to make attachment and calibration easier. Also gather the required hex screwdriver and make sure that the calibration pin is in its storage area on the gripper.
2. CONNECT AND SECURE THE GRIPPER The gantry will move to the front of the robot so you can attach the gripper. Connect the gripper to the extension mount and secure its screws.
3. RUN AUTOMATED CALIBRATION To calibrate the gripper, insert the calibration pin in the front jaw. The gripper will automatically move to touch certain points on the deck and save these calibration values for future use. Then repeat the same process with the calibration pin in the back jaw. Once calibration is complete and you’ve put the pin back in its storage location, the gripper will be ready for use in protocols.
2.5 Relocation
This section provides advice and instructions about how to move your Opentrons Flex robot over short and long distances.

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Short moves
A short move spans a range of distances from “let’s just move it over a little bit” to across the lab, down the hall, or another floor in your building. In these cases, you can move your Flex by hand. Transporting it on a hand cart is also a good option.
Warning: The Flex weighs 88.5 kg. As a result, it requires two people to lift and move it safely.
Reattach the lift handles to move your Flex to a new, nearby location. Lifting and carrying the Flex by its handles is the right way to move the robot short distances. Remove the handles and store them in the User Kit after the move is complete. To prevent damaging the robot, always use the lift handles to pick it up and move it. Do not grab the frame to lift or move your robot.
Long-distance moves
A long-distance move transports your Flex off the grounds of your university, facility, or institution. Across town, to a new city, state, province, or country are all examples of a long-distance move. In this case, you’ll need to pack the Flex to protect it from the elements, shocks, and rough movements that may occur while in transit.
If you’ve kept the shipping crate and internal supports that came with your Flex, you can repackage it in these materials for a long-distance move. Follow the unboxing steps in reverse order to prepare your Flex for a long-distance move. Basically, you should:
Disconnect the power and network cable, if attached. Remove all attached hardware and labware. Reattach the deck plates. Lock the gantry (see the General Moving Advice section below). Remove and store the window panels.
If you kept the original crate:
Reattach the shipping frame to the Flex and secure it to the pallet base using the L-brackets. Add padding and reassemble the shipping crate.

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If you don’t have the original crate and related material, contact a reputable shipping company. They can manage the packing, transportation, and delivery process for you.
General moving advice
DISCONNECT POWER AND NETWORK CABLES Before moving your Flex, don’t forget to: Turn off the power and unplug it from the power supply. Disconnect the Ethernet or USB cable, if used.
LOCK THE GANTRY Before moving your Flex, reinsert the locking screws to hold the gantry in place. The gantry locking points are located: On the left side rail near the front of the robot. Underneath the vertical gantry arm. On the right side rail near the front of the robot. Locking this part of the gantry requires the small
orange bracket and two locking screws.

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HOME THE GANTRY You may not want to lock the gantry if you’re only moving the robot to a nearby location. If you decide not to lock it, at least use the touchscreen or the Opentrons App to send the gantry to its home position before powering it down. To home the gantry via the touchscreen, tap the three-dot menu () and then tap Home gantry. To home the gantry via the Opentrons App: Click Devices. Click on your Flex in the device list. Click the three-dot menu () and then click Home gantry.
REMOVE MODULES In-deck modules and other attachments add extra weight to your Flex. They also affect the robot’s center of gravity, which can make it feel “tippy” when lifting it. To help lighten and balance the robot, remove any attached instruments and labware before you pick it up.
REINSTALL DECK SLOTS We recommend reattaching the deck slots for a long- distance move. Securing the slots in their original locations helps prevent accidental loss. Reattaching the deck slots for short moves around the lab is optional.
POST-MOVE RECALIBRATION You should recalibrate any instruments and modules after reinstalling them. For more details on module calibration, see the Modules chapter.

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Final thoughts about moving
Your Flex is a sturdy and well-built machine, but it is also a precise scientific instrument designed to exacting tolerances. As a result, you should treat it with care when relocating it within your local work area or sending it across the country. This means following the guidance provided here and using your own common sense about how to transport an expensive piece of laboratory equipment. Bottom line: when moving your Flex, err on the side of caution and extra padding.
If you have questions or concerns about relocating your Flex, contact us at [email protected].

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CHAPTER 3
System Description

This chapter describes the hardware systems of Opentrons Flex, which underlie its core lab automation features. The deck, gantry, and instrument mounts of Opentrons Flex enable the use of precision liquidand labware-handling components. The on-device touchscreen enables running protocols and checking on the robot’s status without needing to bring your computer to the lab bench. Wired and wireless connectivity enables additional control from the Opentrons App (see the Software and Operation chapter for more details) and extending the system’s features by attaching peripherals (see the Modules chapter).

3.1 Physical components

Camera

Status Light

Touchscreen

Frame

Gantry Deck
Front Door
Locations of the physical components of Opentrons Flex.

Side Windows Handle Caps

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Frame and enclosure
The frame of the Opentrons Flex robot provides rigidity and structural support for its deck and gantry. All of the mechanical subsystems are situated on and mounted to the main frame. The frame is constructed primarily of sheet metal and aluminum extrusions.
The metal frame has openings for side windows and a front door made of transparent polycarbonate that let you see what’s going on inside Flex. The front door hinges open for access to the interior of the system. With the front door open, you can attach instruments, modules, and deck fixtures; prepare the deck before a protocol; or manipulate the state of the deck during a protocol.
White LED strips on the inside top edges of the frame provide software- controllable ambient lighting. A 2-megapixel camera can photograph the deck and working area for recording and tracking protocol execution.

Deck and working area
The deck is the machined aluminum surface on which automated science protocols are executed. The deck has 12 main ANSI/SLAS-format slots that can be reconfigured to hold labware, modules, and consumables. The deck slots are identified by a coordinate system, with slot A1 at the back left and slot D3 at the front right.

Expansion Slot (for Thermocycler) Working Area

Staging Area

Areas of the deck within Flex.

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The working area is the physical space above the deck that is accessible for pipetting. Labware placed in slots A1 through D3 are in the working area.
Opentrons Flex comes with removable deck slots for all 12 positions in the working area. Each deck slot has corner labware clips for securely placing labware on the deck.
You can reconfigure the deck by replacing slots with other deck fixtures, including the movable trash, waste chute, and module caddies. The expansion slot behind A1 is only used to make additional room for the Thermocycler Module, which occupies slots A1 and B1.
Note: Deck slots are interchangeable within a column (1, 2, or 3) but not across columns; column 1 and column 3 slots are distinct pieces despite their similar size. You can tell which column a slot goes in by orienting the blue labware clip to the back left.
You should leave deck slots installed in locations where you want to place standalone labware. The deck and items placed on it remain static, unless moved by the gripper or manual intervention.
Staging area
The staging area is additional space along the right side of the deck. You can store labware in this location after installing staging area slots. Labware placed in slots A4 through D4 are in the staging area. Flex pipettes cannot reach into the staging area, but the gripper can pick up and move labware to and from this location. Adding extra slots helps keep the working area available for the equipment used in your automated protocols.
Staging area slots are included in certain workstation configurations and are also available for purchase from https://shop.opentrons.com.

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Staging Area with Slots Installed

Deck fixtures
Fixtures are hardware items that replace standard deck slots. They let you customize the deck layout and add functionality to your Flex. Currently, deck fixtures include the staging area slots, the internal trash bin, and the external waste chute. You can only install fixtures in a few specific deck slots. The following table lists the deck locations for each fixture.

Fixture Staging area slots Trash bin Waste chute Waste chute with staging area slot

Slots A3­D3 A1­D1 and A3-D3 D3 only D3 only

Fixtures are unpowered. They do not contain electronic or mechanical components that communicate their current state and deck location to the robot. This means you have to use the deck configuration feature to let the Flex know what fixtures are attached to the deck and where they’re located.

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You can access the deck configuration settings from the touchscreen via the three-dot () menu and from the Opentrons App. See the Deck Configuration section of the Software and Operation chapter for information on how to configure the deck from the touchscreen.

Waste chute
The Opentrons Flex Waste Chute transfers liquids, tips, tip racks, and well plates from the Flex enclosure to a trash receptacle placed below its external opening. The waste chute attaches to a deck plate adapter that fits in slot D3. It also comes with a special window half panel that lets the chute extend out of the front of the robot.
Components of the waste chute.

Cover Deck Plate Adapter

Waste Chute
Deck Plate Adapter with Staging Area

Staging area slots
Staging area slots are ANSI/SLAS compatible deck pieces that replace standard slots in column 3 and add new slots to the staging area — all without losing space in the working area. You can install a single slot or a maximum of four slots to create a new column (A4 to D4) along the right side of the deck. Note, however, that replacing deck slot A3 requires moving the trash bin. By adding staging area slots to the deck, your Flex robot can store more labware and operate more efficiently.

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Flex staging area slot.
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SLOT INSTALLATION
To install, remove the screws that attach a standard slot to the deck and replace it with the staging area slot. After installation, use the touchscreen or Opentrons App to tell the robot you’ve added a staging area slot to the deck.
Installing a staging area slot.

SLOT COMPATIBILITY Staging area slots are compatible with the Flex instruments, modules, and labware listed below.

Flex component Gripper Pipettes Modules
Labware

Staging area compatibility
The Flex Gripper can move labware to or from staging area slots.
Flex pipettes cannot reach the staging area. Use the gripper to move tip racks and labware from the staging area to the working area before pipetting.
The Magnetic Block GEN1 can be placed in column 3 on top of a staging area slot. Modules are not supported in column 4.
Powered modules such as the Heater-Shaker and Temperature Module fit into caddies that can be placed in column 3. You can’t add a staging area slot to a position occupied by a module caddy.
Staging area slots have the same ANSI/SLAS dimensions as standard deck slots. Use gripper-compatible labware in the staging area, or manually add and remove labware from this location.

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Movement system
Attached to the frame is the gantry, which is the robot’s movement and positioning system. The gantry moves separately along the x- and y-axis to position the pipettes and gripper at precise locations for protocol execution. Movement along these axes is precise to the nearest 0.1 mm. The gantry is controlled by 36 VDC hybrid bipolar stepper motors. In turn, attached to the gantry are the pipette mounts and the extension mount. These move along the z-axis to position the pipettes and gripper at precise locations for protocol execution. Movement along this axis is controlled by 36 VDC hybrid bipolar stepper motors. The electronics contained in the gantry provide 36 VDC power and communications to the pipettes and gripper, when attached.
Gantry

Pipette Mounts

Extension Mount

Location of instrument mounts on Flex.

Touchscreen and LED displays
The primary user interface is the 7-inch LCD touchscreen, located on the front right of the robot. The touchscreen is covered with Gorilla Glass 3 for scratch and damage resistance. Access many features of Flex right on the touchscreen, including:

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Protocol management Protocol setup, execution, and monitoring Labware management Robot settings System software and firmware updates Operation logs and error notifications
For more information on using Flex via the touchscreen, see the Touchscreen Operation section of the Software and Operation chapter.
The status light is a strip of LEDs along the top front of the robot that provides at-a-glance information about the robot. Different colors and patterns of illumination can communicate various success, failure, or idle states:

LED color White Neutral states
Green Normal states
Blue Mandatory states Yellow Abnormal states Red Emergency states

LED pattern Solid Pulsing
Blinks twice
Solid Pulsing Pulsing

Robot status
Powered on and not running a protocol Robot is busy (e.g., updating software or firmware, setting up protocol run, canceling protocol run) Action is complete (e.g., protocol stored, software updated, instrument attached or detached) Protocol is running Protocol is complete Protocol is paused

Solid

Software error

Blinks three times, repeatedly

Physical error (e.g., instrument crash)

The status light can also be disabled in the robot settings.

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3.2 Pipettes
Opentrons pipettes are configurable devices used to move liquids throughout the working area during the execution of protocols. There are several Opentrons Flex pipettes, which can handle volumes from 1 µL to 1000 µL in 1, 8, or 96 channels:
Opentrons Flex 1-Channel Pipette (1­50 µL) Opentrons Flex 1-Channel Pipette (5­1000 µL) Opentrons Flex 8-Channel Pipette (1­50 µL) Opentrons Flex 8-Channel Pipette (5­1000 µL) Opentrons Flex 96-Channel Pipette (5­1000 µL)
Pipettes attach to the gantry using captive screws on the front of the pipette. 1-channel and 8-channel pipettes each occupy one pipette mount (left or right); the 96-channel pipette occupies both mounts. For details on installing pipettes, see Instrument Installation and Calibration.

Captive Attachment
Screws

Captive Attachment Screws

Ejector

Nozzles (Replaceable O-rings)

Nozzles (Fixed O-rings)

Locations of components of the 1-, 8-, and 96-channel pipettes.

Ejector

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The pipettes pick up disposable plastic tips by pressing them onto the pipette nozzles, and then use the tips to aspirate and dispense liquids. The amount of total force required for pickup increases as more tips get picked up simultaneously. For smaller numbers of tips, the pipette attaches tips by pushing each pipette nozzle down into a tip. To achieve the necessary force to pick up a full rack of tips, the 96-channel pipette also pulls the tips upward onto the nozzles. This pulling action requires placing tip racks into a tip rack adapter, rather than directly in a deck slot. To discard tips (or return them to their rack), the pipette ejector mechanism pushes the tips off of the nozzles.

Pipette specifications
Opentrons Flex pipettes are designed to handle a wide range of volumes. Because of their wide overall range, they can use multiple sizes of tips, which affect their liquid-handling characteristics. Opentrons has tested Flex pipettes for accuracy and precision in a number of tip and liquid volume combinations:

Pipette
Flex 1-Channel
50 µL
Flex 1-Channel
1000 µL
Flex 8-Channel
50 µL
Flex 8-Channel
1000 µL

Tip Capacity 50 µL 50 µL 50 µL 50 µL 50 µL 200 µL 1000 µL 50 µL 50 µL 50 µL 50 µL 50 µL 200 µL 1000 µL

Tested Volume 1 µL 10 µL 50 µL 5 µL 50 µL
200 µL 1000 µL
1 µL 10 µL 50 µL 5 µL 50 µL 200 µL 1000 µL

Accuracy %D 8.00% 1.50% 1.25% 5.00% 0.50% 0.50% 0.50% 10.00% 2.50% 1.25% 8.00% 2.50% 1.00% 0.70%

Precision %CV 7.00% 0.50% 0.40% 2.50% 0.30% 0.15% 0.15% 8.00% 1.00% 0.60% 4.00% 0.60% 0.25% 0.15%

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Flex 96-Channel
1000 µL

50 µL 50 µL 200 µL 1000 µL

5 µL 50 µL 200 µL 1000 µL

10.00% 2.50% 1.50% 1.50%

5.00% 1.25% 1.25% 1.50%

Keep this accuracy information in mind when choosing tips for your pipette. In general, for best results you should use the smallest tips that meet the needs of your protocol.

Note: Opentrons performs volumetric testing of Flex pipettes to ensure that they meet the accuracy and precision specifications listed above. You do not have to calibrate the volume that your pipettes dispense before use. You only have to perform positional calibration. See the next section, as well as the Pipette Installation section of the Installation and Relocation chapter, for details.
The Opentrons Care and Opentrons Care Plus services include yearly pipette replacement and certificates of calibration. See the Servicing Flex section of the Maintenance and Service chapter for details.

Pipette calibration
The User Kit includes a metal pipette calibration probe, which you use during positional calibration. During protocol runs, safely store the probe on the magnetic holder on the front pillar of the robot. During the calibration process, attach the probe to the appropriate nozzle and lock it in place. The robot moves the probe to calibration points on the deck to measure the pipette’s exact position.
Pipette tip rack adapter
The Opentrons Flex 96-channel pipette ships with four tip rack adapters. These are precision formed aluminum brackets that you place on the deck. The adapters hold Flex 50 L, 200 L, and 1000 µL tip racks.

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Because of the force involved, the 96-channel pipette requires an adapter to attach a full tip rack properly. During the attachment procedure, the pipette moves over the adapter, lowers itself onto the mounting pins, and pulls tips onto the pipettes by lifting the adapter and tip rack. Pulling the tips, rather than pushing, provides the leverage needed to secure tips to the pipettes and prevents warping the deck surface. When finished, the 96-channel pipette lowers the adapter and empty tip rack onto the deck. See the Tips and tip racks section of the Labware chapter for more information.
Partial tip pickup
The 96-channel pipette can pick up a full rack of tips or a smaller number of tips. This increases the number of applications you can perform with the 96-channel pipette, since it occupies both pipette mounts.
Currently, the 96-channel pipette supports partial tip pickup for 8 tips in a column layout. In this configuration, the pipette either uses its leftmost nozzles to pick up tips right-to-left from a tip rack, or its rightmost nozzles to pick up tips left-to-right from a tip rack.
When picking up fewer than 96 tips from a tip rack, the rack must be placed directly on the deck, not in the tip rack adapter.
Pipette sensors
Opentrons Flex pipettes have a number of sensors that detect and record data about the status of the pipette and any tips it has picked up.
CAPACITANCE SENSORS
In combination with a metal probe or conductive tip, the capacitance sensors detect when the pipette makes contact with something. Detection of contact between the metal probe and the deck is used in the automated pipette calibration and module calibration processes.
1-channel pipettes have one capacitance sensor, while multi-channel pipettes have two: on channels 1 and 8 of 8-channel pipettes, and on channels 1 and 96 (positions A1 and H12) of the 96-channel pipette.

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OPTICAL TIP PRESENCE SENSORS
A photointerruptor switch detects the position of the pipette’s tip ejector mechanism, confirming whether tips were successfully picked up or dropped. 1-channel, 8-channel, and 96-channel pipettes all have a single optical sensor that monitors tip attachment across all channels.
Pipette firmware updates
Opentrons Flex automatically updates pipette firmware to keep it in sync with the robot software version. Pipette firmware updates are typically quick, and occur whenever:
You attach a pipette. The robot restarts.
If, for any reason, your pipette firmware and robot software versions get out of sync, you can manually update the firmware in the Opentrons App.
1. Click Devices. 2. Click on your Flex in the device list. 3. Under Instruments and Modules, the out-of-sync pipette will show a warning banner reading
“Firmware update available.” Click Update now to begin the update.
You can view the currently installed firmware version of any attached pipette. On the touchscreen, go to Instruments and tap the pipette name. In the Opentrons App, find the pipette card under Instruments and Modules, click the three-dot menu (), and then click About pipette.

3.3 Gripper
The gripper moves labware throughout the working area and staging area during the execution of protocols. The gripper attaches to the extension mount, which is separate from the pipette mounts; the gripper can be used with any pipette configuration. For details on installing the gripper, see Instrument Installation and Calibration.
The gripper can move labware across the deck and onto or off of modules. The gripper can manipulate certain fully skirted well plates, deep well plates, and tip racks. For more details on what labware the gripper can move, see the Labware and the Opentrons Flex Gripper section of the Labware chapter, or consult the Opentrons Labware Library.

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Gripper specifications
The jaws perform the primary motion of the gripper, which is to open or close two parallel paddles to apply or release force on the sides of labware. Movement of the jaws is controlled by a 36 VDC brushed motor connected to a rack-and-pinion gear system.
To move a piece of labware that has been gripped by the jaws, the gantry lifts the gripper along the z-axis, moves it laterally, and then lowers it into the labware’s new position.
Locations of components of the gripper.

Attachment screws
Calibration pin Jaws Paddles

Gripper calibration
The gripper includes a metal calibration pin. The calibration pin is located in a recessed storage area on the lower part of the gripper. A magnet holds the pin in place. To remove the calibration pin, grasp it with your fingers and pull gently. To replace the pin, put it back in the storage slot. You’ll know it’s secure when it snaps into place.
When calibrating the gripper, attach the pin to each jaw in turn. The robot moves the pin to calibration points on the deck to measure the gripper’s exact position.
During protocol runs, place the pin in its storage area for safekeeping. Contact us at [email protected] if you lose the calibration pin.

Gripper firmware updates
Opentrons Flex automatically updates the gripper firmware to keep it in sync with the robot software version. Gripper firmware updates are typically quick, and occur whenever:

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You attach the gripper. The robot restarts.
If, for any reason, your gripper firmware and robot software versions get out of sync, you can manually update the firmware in the Opentrons App.
1. Click Devices. 2. Click on your Flex in the device list. 3. Under Instruments and Modules, the out-of-sync gripper will show a warning banner reading
“Firmware update available.” Click Update now to begin the update.
You can view the currently installed firmware version of the gripper. On the touchscreen, go to Instruments and tap the gripper. In the Opentrons App, find the gripper card under Instruments and Modules, click the three-dot menu (), and then click About gripper.
3.4 Emergency Stop Pendant
The Emergency Stop Pendant (E-stop) is a dedicated hardware button for quickly stopping robot motion. Opentrons Flex requires a functional, disengaged E-stop to be attached at all times. When you press the stop button, Flex cancels any running protocol or setup workflow as quickly as possible and prevents most robot motion.
When to use the E-stop
You may need to press the E-stop:
When there is imminent risk of injury or harm to a user. When there is imminent risk of damage to the robot or other hardware. When samples or reagents are in imminent danger of contamination. After a hardware collision.
Ideally you should never have to press the E-stop (except during infrequent hardware quality testing).
Do not use the E-stop to cancel normal, expected operations. Instead, use the software button on the touchscreen or in the Opentrons App. Pausing via software will let you resume or cancel your protocol, whereas pressing the E-stop always cancels the protocol immediately.

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Engaging and releasing the E-stop
The E-stop has a press-to-engage, twist-to-release mechanism.
Engage: Push down firmly on the red button. Flex will enter the stopped state. Resolve: Once stopped, safely address any problems in the working area, such as clearing spills,
removing labware, or moving the gantry (it should move freely and easily by hand). Release: Twist the button clockwise. It will pop up to its disengaged position. Reset: On the touchscreen or in the Opentrons App, confirm that you are ready for Flex to resume
motion. The gantry will return to its home position and module activity will resume.
In the stopped state, Flex and connected hardware will behave as follows:

Hardware Gantry Pipettes
Gripper
Heater-Shaker Module
Temperature Module Thermocycler Module Status light Touchscreen

Behavior
Automated horizontal motion is halted. Manual horizontal motion is allowed.
Vertical motion of pipettes is halted. The motor brakes on vertical axes are engaged to prevent
pipettes from falling. Plunger motion and tip pickup is halted.
Vertical motion of the gripper is halted. The motor brake on the vertical axis is engaged to prevent the
gripper from falling. The jaw motors that exert gripping force remain enabled, so
the gripper will not drop labware it may be carrying.
The shaker stops and homes. The labware latch opens. Heating is disabled.
Heating or cooling is disabled.
Heating or cooling is disabled.
The light turns red.
A cancellation message takes over the screen. An on-screen indicator shows when you have successfully
disengaged the stop button.

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3.5 Connections

On/Off Switch

Side Covers

USB-A Ports

IEC Power Inlet

Ports AUX-1, AUX-2, USB-B, Ethernet

Power connection
Opentrons Flex connects to a power source via a standard IEC-C14 inlet. The robot contains an internal full-range AC/DC power supply, accepting 100­240 VAC, 50/60 Hz input and converting it to 36 VDC. All other internal electronics are powered by the 36 VDC supply.

Warning: Only use the power cord provided with the robot. Do not use a power cord with inadequate current or voltage ratings.
Keep the power cord free of obstructions so you can remove it if necessary.

There is also a CR1220 coin cell battery to power the robot’s real-time clock when not connected to mains power. The battery is located inside the touchscreen enclosure. Contact Opentrons Support for more information if you think you need to replace the battery.

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USB and auxiliary connections
Opentrons Flex has 10 total USB ports located in different areas of the robot, which serve different purposes.
The 8 rear USB-A ports (numbered USB-1 through USB-8) and 2 auxiliary ports (M12 connectors numbered AUX-1 and AUX-2) are for connecting Opentrons modules and accessories. See the Modules chapter for more information on connecting these devices and using them in your protocols. The rear USB-B port is for connecting the robot to a laptop or desktop computer, to establish communication with the Opentrons App running on the connected computer. The front USB-A port (USB-9), located below the touchscreen display, has the same functionality as the rear USB-A ports.
Note: The USB ports are power-limited to protect the robot and connected devices. Power delivery is split internally into three port groups: the left rear USB-A ports (USB-1 through USB-4), the right rear USB-A ports (USB-5 through USB-8), and the front USB-A port. Each of these groups will deliver a maximum of 500 mA to connected USB 2.0­compatible devices.
Network connections
Opentrons Flex can connect to a local area network through a wired (Ethernet) or wireless (Wi-Fi) connection.
The Ethernet port is located on the rear of the robot. Connect it to an Ethernet hub or switch on your network. Or, starting in robot system version 7.1.0, connect it directly to an Ethernet port on your computer. The internal Wi-Fi module supports 802.11 ac/a/b/g/n networks with a dual-band 2.4/5 GHz antenna.

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3.6 System specifications

General specifications

Dimensions Weight Deck slots
Touchscreen
Wi-Fi Ethernet USB
Camera Robot power input
Mains supply voltage fluctuation Mains supply frequency fluctuation Distribution system Short-circuit supply current Frame composition Window composition Ventilation requirements

87 × 69 × 84 cm / 34.25 × 27 × 33 in (W, D, H)
88.5 kg / 195 lb 12 ANSI/SLAS-compatible slots in working area
(accessible to pipettes) 4 additional slots for staging tips and labware
(accessible only to gripper) 7-inch LCD touchscreen with scratch- and damage- resistant Gorilla Glass 3
802.11 ac/a/b/g/n dual-band (2.4/5 GHz)
100 Mbps 9 USB-A ports 1 USB-B port USB 2.0 speed
2MP, photo and video 100­240 VAC, 50­60 Hz, 1 4.0 A/115 VAC, 2.0 A/230 VAC
±10%
±5%
TN-S
6.3 A
Rigid steel and CNC aluminum design
Removable polycarbonate side windows and front door At least 20 cm / 8 in between the unit and a wall

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Connected PC requirements

The Opentrons App runs on: Windows 10 or later macOS 10.10 or later Ubuntu 12.04 or later

Environmental specifications

Environmental conditions Ambient temperature Relative humidity Pollution degree

Indoor use only +20 to +25 °C (recommended) 40­60%, non-condensing (recommended) 2 (non-conductive pollution only)

For additional information on acceptable environmental conditions for use and transport, see the Environmental Conditions section of the Installation and Relocation chapter.

Certifications

Certifications complete Not certified/validated

CE, ETL, FCC, ISO 9001 IVD, GMP

A summary of certification information is printed on a sticker on the back of Flex, near the on/off switch. For detailed certification and compliance information, see the Regulatory Compliance section in the Introduction.

Serial number
Every Flex has a unique serial number. The format of the serial number provides additional information, including the robot’s date of production. For example, the serial number FLXA1020231007001 would indicate:

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Characters FLX A10 2023 10 07 001

Category Model Version Year Month Day Unit

Meaning The robot is an Opentrons Flex. A code for the production version of the robot. The robot was made in 2023. The robot was made in October. The robot was made on the 7th day of the month. A unique number for robots made on a certain day.

You can find the serial number for your Flex:
On the certification sticker on the back of Flex, near the on/off switch. On the reverse side of the touchscreen (towards the working area). In the Opentrons App under Devices > your Flex > Robot settings > Advanced.

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Modules
Opentrons Flex integrates with a number of Opentrons hardware modules. All modules are peripherals that occupy deck slots, and most are controlled by the robot over a USB connection.
This chapter describes the functions and physical specifications of modules that are compatible with the Opentrons Flex system, as well as how to attach and calibrate them. For further details on module setup and use, consult the manuals for the individual modules. For details on integrating modules into your protocols, see the Protocol Designer section of the Protocol Development chapter or the online Python Protocol API documentation.
4.1 Supported modules
Opentrons Flex is compatible with four types of on-deck Opentrons modules:
The Heater-Shaker Module provides on-deck heating and orbital shaking. The module can be heated to 95 °C, and can shake samples from 200 to 3000 rpm.
The Magnetic Block is a passive device that holds labware close to its high- strength neodymium magnets. The OT-2 Magnetic Module GEN1 and GEN2, which actively move their magnets up and down relative to labware, are not supported on Opentrons Flex.
The Temperature Module is a hot and cold plate module that is able to maintain steady state temperatures between 4 and 95 °C.
The Thermocycler Module provides on-deck, fully automated thermocycling, enabling automation of upstream and downstream workflow steps. Thermocycler GEN2 is fully compatible with the gripper. Thermocycler GEN1 cannot be used with the gripper, and is therefore not supported on Opentrons Flex.
Some modules originally designed for the OT-2 are compatible with Flex, as summarized in the table below. A checkmark indicates compatibility, and an X indicates incompatibility.

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Device type and generation Heater-Shaker Module GEN1 Magnetic Module GEN1 Magnetic Module GEN2 Magnetic Block GEN1 Temperature Module GEN1 Temperature Module GEN2 Thermocycler Module GEN1 Thermocycler Module GEN2 HEPA Module

OT-2

Flex

×

×

×

×

×

×

4.2 Module caddy system
Compatible modules are designed to fit into caddies that occupy space below the deck. This system allows labware on top of modules to remain closer to the deck surface, and it also allows for below-deck cable routing so the deck stays tidy during your protocol runs.

Caddies for the Heater-Shaker, Temperature, and Thermocycler Modules.

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To fit a module into the deck surface, it must first be placed into the corresponding module caddy. Each type of compatible module has its own caddy design that aligns the module and labware precisely with the surrounding deck. (The exception is the Magnetic Block, which does not require power or USB cable routing and thus sits directly on the deck surface.) Caddies for modules that occupy a single slot can be placed anywhere in column 1 or 3; the Thermocycler can only be placed in slots A1 and B1 simultaneously.
In general, to install a module caddy:
1. Remove any deck slots from the location where the module will go. 2. Seat the module into its caddy and tighten its anchors. 3. Route the module power and USB cables through the side covers, up through the empty deck slot, and
attach them to the module. 4. Seat the module caddy into the slot and screw it into place.
For exact installation instructions, consult the Quickstart Guide or Instruction Manual for the specific module. Cable connections and method of attachment to the caddy vary by module.
4.3 Module calibration
When you first install a module on Flex, you need to run automated positional calibration. This process is similar to positional calibration for instruments, and ensures that Flex moves to the exact correct locations for optimal protocol performance. During calibration, Flex will move to locations on a module calibration adapter, which looks similar to the calibration squares that are part of removable deck slots.

Calibration adapters for the Heater-Shaker, Temperature, and Thermocycler Modules.
Module calibration is required for all modules that install via a caddy: the Heater-Shaker, Temperature, and Thermocycler Modules. The Magnetic Block doesn’t require calibration, and is ready for use as soon as you place it on the deck.

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When to calibrate modules
Flex automatically prompts you to perform calibration when you connect and power on a module that doesn’t have any stored calibration data. (You can dismiss this prompt, but you won’t be able to run protocols with the module until you calibrate it.)
Once you’ve completed calibration, Flex stores the calibration data and module serial number for future use. Flex won’t prompt you to recalibrate unless you delete the calibration data for that module in the robot settings. You can freely power your module on and off, or even move it to another deck slot, without needing to recalibrate. If you want to recalibrate, you can begin the process at any time from the module card in the Opentrons App. (Recalibration is not available from the touchscreen.)
How to calibrate modules
Instructions on the touchscreen or in the Opentrons App will guide you through the calibration procedure. In general the steps are:
1. Gather the required equipment, including the module calibration adapter and pipette calibration probe. 2. Place the calibration adapter on the module surface and ensure that it is completely level.
Some modules may require you to fasten the adapter to the module. 3. Attach the calibration probe to a pipette. 4. Flex will automatically move to touch certain points on the calibration adapter and save these
calibration values for future use.
Once calibration is complete and you’ve removed the adapter and probe, the module will be ready for use in protocols.
At any time, you can view and manage your module calibration data in the Opentrons App. Go to Robot Settings for your Flex and click on the Calibration tab.

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4.4 Heater-Shaker Module GEN1

Heater-Shaker features
HEATING AND SHAKING
The Heater-Shaker provides on-deck heating and orbital shaking. The module can be heated to 95 °C, with the following temperature profile:
Temperature range: 37­95 °C Temperature accuracy: ±0.5 °C at 55 °C Temperature uniformity: ±0.5 °C at 55 °C Ramp rate: 10 °C/min
The module can shake samples from 200 to 3000 rpm, with the following shaking profile:
Orbital diameter: 2.0 mm Orbital direction: Clockwise Speed range: 200­3000 rpm Speed accuracy: ±25 rpm
The module has a powered labware latch for securing plates to the module prior to shaking.

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THERMAL ADAPTERS A compatible thermal adapter is required for adding labware to the Heater-Shaker. Adapters can be purchased directly from Opentrons at https://shop.opentrons.com. Currently available Thermal Adapters include:

Universal Flat Adapter

PCR Adapter

Deep Well Adapter

96 Flat Bottom Adapter

SOFTWARE CONTROL
The Heater-Shaker is fully programmable in Protocol Designer and the Python Protocol API. The Python API additionally allows for other protocol steps to be performed in parallel while the Heater-Shaker is active. See Non-blocking commands in the API documentation for details on adding parallel steps to your protocols.
Outside of protocols, the Opentrons App can display the current status of the Heater-Shaker and can directly control the heater, shaker, and labware latch.

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Heater-Shaker specifications

Dimensions Weight Module power input Power adapter input Mains supply voltage fluctuation Overvoltage Power consumption

152 × 90 × 82 mm (L/W/H) 1.34 kg 36 VDC, 6.1 A 100­240 VAC, 50/60 Hz ±10% Category II Idle: 3 W

Typical: Shaking: 4­11 W Heating: 10­30 W Heating and shaking: 10­40 W

Environmental conditions Ambient temperature Relative humidity Altitude Pollution degree

Maximum: 125­130 W Indoor use only 20­25 °C Up to 80%, non-condensing Up to 2,000 m above sea level 2

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4.5 Magnetic Block GEN1

Magnetic Block features
The Opentrons Magnetic Block GEN1 is a magnetic 96-well plate holder. Magnetic blocks are used in protocols that rely on magnetism to pull particles out of suspension and retain them in well plates during wash, rinse, or other elution procedures. For example, automated NGS preparation; purifying genomic and mitochondrial DNA, RNA, or proteins; and other extraction procedures are all use cases that can involve magnetic blocks.
MAGNETIC COMPONENTS
The Magnetic Block is unpowered, does not contain any electronic components, and does not move magnetic beads up or down in solution. The wells consist of 96 high-strength neodymium ring magnets fixed to a spring-loaded bed, which helps maintain tolerances between the block and pipettes while running automated protocols.
SOFTWARE CONTROL
The Magnetic Block GEN1 is fully programmable in Protocol Designer and the Python Protocol API.

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Outside of protocols, however, the touchscreen and the Opentrons App are not aware of and cannot display the current status of the Magnetic Block GEN1. This is an unpowered module. It does not contain electronic or mechanical components that can communicate with the Flex robot. You “control” the Magnetic Block via protocols that use the Opentrons Flex Gripper to add and remove labware from this module.

Magnetic Block specifications

Dimensions Weight Module power Magnet grade Environmental conditions Ambient temperature Relative humidity Altitude Pollution degree

136 × 94 × 45 mm (L/W/H) 1.13 kg None, module is unpowered N52 neodymium Indoor use only 20­25 °C 30­80%, non-condensing Up to 2000 m above sea level 2

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4.6 Temperature Module GEN2

Temperature Module features
HEATING AND COOLING
The Opentrons Temperature Module GEN2 is a hot and cold plate module. It is often used in protocols that require heating, cooling, or temperature changes. The module can reach and maintain temperatures ranging from 4 °C to 95 °C within minutes, depending on the module’s configuration and contents.
THERMAL BLOCKS
To hold labware at temperature, the module uses aluminum thermal blocks. The module comes with 24well and 96-well thermal blocks. The Temperature Module caddy comes with a deep well block and a flat bottom block designed for use with the Flex Gripper. The blocks hold 1.5 mL and 2.0 mL tubes, 96-well PCR plates, PCR strips, deep well plates, and flat bottom plates.

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Note: The module also ships with a flat bottom block for the OT-2. Do not use the OT-2 block with Flex. The flat bottom block for Flex has the words “Opentrons Flex” on its top surface. The one for OT-2 does not.

24-well thermal block

96-well thermal block

Deep well thermal block

Flat bottom thermal block for Flex

WATER BATHS AND HEATING
Because air is a good thermal insulator, gaps between labware and a thermal block can affect the time-totemperature performance of the Temperature Module. Placing a little water in the 24- or 96-well thermal blocks eliminates air gaps and improves heating efficiency. The ideal amount of water depends on the thermal block and labware. See the Temperature Module White Paper for further recommendations.

SOFTWARE CONTROL
The Temperature Module is fully programmable in Protocol Designer and the Python Protocol API.
Outside of protocols, the Opentrons App can display the current status of the Temperature Module and can directly control the temperature of the surface plate.

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Temperature Module specifications

Dimensions Weight Module power
Environmental conditions Ambient temperature Relative humidity Altitude Pollution degree

194 × 90 × 84 mm (L/W/H) 1.5 kg Input: 100­240 VAC, 50/60 Hz, 4.0 A Output: 36 VDC, 6.1 A, 219.6 W max Indoor use only <22 °C (recommended for optimal cooling) Up to 60%, non-condensing Up to 2000 m above sea level 2

4.7 Thermocycler Module GEN2

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Thermocycler features
The Opentrons Thermocycler Module GEN2 is a fully automated on-deck thermocycler, providing hands-free PCR in a 96-well plate format. Its heated lid and disposable seal fit tightly over the plate, ensuring efficient sample heating and minimal evaporation.
HEATING AND COOLING The Thermocycler’s block can heat and cool, and its lid can heat, with the following temperature profile: Thermal block temperature range: 4­99 °C Thermal block maximum heating ramp rate: 4.25 °C/s from GEN2 ambient to 95 °C Thermal block maximum cooling ramp rate: 2.0 °C/s from 95 °C to ambient Lid temperature range: 37­110 °C Lid temperature accuracy: ±1 °C The automated lid can be opened or closed as needed during protocol execution.
THERMOCYCLER PROFILES The Thermocycler can execute profiles: automatically cycling through a sequence of block temperatures to perform heat-sensitive reactions.
RUBBER AUTOMATION SEALS The Thermocycler comes with rubber automation seals to help reduce evaporation. Each seal must be sterilized before use and can be used for several runs. Additional seals can be purchased directly from Opentrons at https://shop.opentrons.com.
SOFTWARE CONTROL The Thermocycler is fully programmable in Protocol Designer and the Python Protocol API. Outside of protocols, the Opentrons App can display the current status of the Thermocycler and can directly control the block temperature, lid temperature, and lid position.

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Thermocycler specifications

Dimensions (lid open) Dimensions (lid closed) Weight (including rear duct) Power adapter voltage Power adapter current Overvoltage Environmental conditions Ambient temperature Relative humidity Altitude Ventilation requirements

244.95 × 172 × 310.1 mm (L/W/H) 244.95 × 172 × 170.35 mm (L/W/H) 8.4 kg 100­240 V at 50/60 Hz 8.5­5 A Category II Indoor use only 20­25 °C (ideal); 2­40 °C (acceptable) 30­80%, non-condensing Up to 2000 m above sea level At least 20 cm / 8 in between the unit and a wall

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CHAPTER 5
Labware
This chapter covers items in the Opentrons Labware Library you can use with Opentrons Flex and the Opentrons Flex Gripper. It also covers custom labware and, for our power users, links labware components to their corresponding JSON file definitions.
You can purchase labware from the original equipment manufacturers or from the Opentrons shop at https://shop.opentrons.com. And, Opentrons is always working to verify new labware definitions. See the Labware Library (linked above) for the latest listings.
5.1 Labware concepts
Labware encompasses more than just the objects placed on the deck and used in a protocol. To help you understand Opentrons labware, let’s examine this topic from three different perspectives. For the Opentrons Flex, labware includes items in our Labware Library, data that defines each piece of labware, and custom labware.
Labware as hardware
The Labware Library includes everything you can use by default with Opentrons Flex. These are durable components and consumable items that you work with, reuse, or discard while running a protocol. You don’t need to take any special steps to work with the items in the Labware Library. The Flex robot knows how to work with everything in the library automatically.
Labware as data
Labware information is stored in Javascript object notation (JSON) files with .json file extensions. A JSON file includes spatial dimensions (length, width, height), volumetric capacity (L, mL), and other metrics that define surface features, their shapes, and locations. When running a protocol, the Flex reads these .json files to know what labware is on the deck and how to work with it.

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Custom labware
Custom labware is labware that is not included in the Labware Library or is labware created by the Custom Labware Creator. However, sometimes the idea of custom labware comes burdened by notions of complexity, expense, or difficulty. But, custom labware shouldn’t be hard to understand or create. Let’s take a moment to unpack the concept of custom labware.
As an example, the Opentrons Labware Library includes 96-well plates (200 L) from Corning and BioRad, but other manufacturers make these well plates too. And, thanks to commonly accepted industry standards, the differences among these ubiquitous lab items are minor. However, an ordinary 200 L, 96-well plate from Stellar Scientific, Oxford Lab, or Krackeler Scientific (or any other supplier for that matter) is “custom labware” for the Flex because it isn’t pre-defined in our Labware Library. Additionally, minor differences in labware dimensions can have a drastic impact on the success of your protocol run. For this reason, it’s important to have an accurate labware definition for each labware you want to use in your protocol.
Also, while custom labware could be an esoteric, one-off piece of kit, most of the time it’s just the tips, plates, tubes, and racks used every day in labs all over the world. Again, the only difference between Opentrons labware and custom labware is the new item is not predefined in the software that powers the robot. The Flex can, and does, work with other basic labware items or something unique, but you need to record that item’s characteristics in a labware definition JSON file and import that data into the Opentrons App. See the Custom Labware Definitions section below for more information.
To sum up, labware includes:
Everything in the Opentrons Labware Library. Labware definitions: data in a JSON file that defines shapes, sizes, and capabilities of individual items
like well plates, tips, reservoirs, etc. Custom labware, which are items that aren’t included in the Labware Library.
After reviewing these important concepts, let’s examine the categories and items in the Opentrons Labware Library. After that, we’ll finish the chapter with an overview of the data components of a labware file and summarize the Opentrons features and services that help you create custom labware.

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5.2 Reservoirs
The Opentrons Flex works by default with the single- and multi-well reservoirs listed below. Using these reservoirs helps reduce your prep work burden because they’re automation-ready right out of the box. Reservoir information is also available in the Opentrons Labware Library.

Single-well reservoirs

Manufacturer Specifications

Agilent

290 mL V bottom

Axygen

90 mL Flat bottom

NEST

195 mL Flat bottom

290 mL V bottom

API load name
agilient1 reservoir_290ml
axygen1 reservoir_90ml
nest1 reservoir_195ml
nest1 reservoir_290ml

Multi-well reservoirs

Manufacturer Specifications

NEST

12 wells 15 mL/well V bottom

USA Scientific

12 wells 22 mL/well V bottom

API load name nest12 reservoir_15ml
usascientific12 reservoir_22ml

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Reservoirs and API definitions
The Opentrons Labware Library defines the characteristics of the reservoirs listed above in separate JSON files. The robot and the Opentrons Python API rely on these JSON definitions to work with labware used by your protocols. For example, when working with the API, the ProtocolContext.load_labware function accepts these labware names as valid parameters in your code. Linked API load names connect to the reservoir labware definitions in the Opentrons GitHub repository.
Custom reservoir labware
Try creating a custom labware definition with the Opentrons Labware Creator if a reservoir you’d like to use isn’t listed here. A custom definition combines all the dimensions, metadata, shapes, volumetric capacity, and other information in a JSON file. The Opentrons Flex needs this information to understand how to work with your custom labware. See the Custom Labware Definitions section section for more information.
5.3 Well plates
The Opentrons Flex works by default with well plates listed below. Using these well plates helps reduce your prep work burden because they’re automation- ready right out of the box. Well plate information is also available in the Opentrons Labware Library.

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6-well plates
Manufacturer Corning

Specifications
6 wells 16.8 mL/well Circular wells, flat bottom

API load name corning_6_wellplate_16.8ml_flat

12-well plates
Manufacturer Corning

Specifications
12 wells 6.9 mL/well Circular wells, flat bottom

API load name corning_12_wellplate_6.9ml_flat

24-well plates
Manufacturer Corning

Specifications
24 wells 3.4 mL/well Circular wells, flat bottom

API load name corning_24_wellplate_3.4ml_flat

48-well plates
Manufacturer Corning

Specifications
48 wells 1.6 mL/well Circular wells, flat bottom

API load name corning_48_wellplate_1.6ml_flat

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96-well plates
Manufacturer Bio-Rad Corning NEST
Opentrons Thermo Scientific
USA Scientific

Specifications
96 wells 200 µL/well Circular wells, V bottom
96 wells 360 µL/well Circular wells, flat bottom
96 wells 100 µL/well Circular wells, V bottom PCR full skirt
96 wells 200 µL/well Circular wells, flat bottom
96 deep wells 2000 µL/well Square wells, V bottom
Tough 96 wells 200 µL/well Circular wells, V bottom PCR full skirt
Nunc 96 deep wells 1300 µL/well Circular wells, U bottom
Nunc 96 deep wells 2000 µL/well Circular wells, U bottom
96 deep wells 2.4 mL/well Square wells, U bottom

API load name biorad_96_wellplate_200ul_pcr
corning_96_wellplate_360ul_flat
nest_96_wellplate_100ul_pcr_full_skirt
nest_96_wellplate_200ul_flat
nest_96_wellplate_2ml_deep
opentrons_96_wellplate_200ul_pcrfull skirt
thermoscientificnunc_96wellplate 1300ul thermoscientificnunc_96wellplate 2000ul usascientific_96_wellplate_2.4ml_deep

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384-well plates

Manufacturer Applied Biosystems Bio-Rad
Corning

Specifications
384 wells 40 µL/well Circular wells, V bottom
384 wells 50 µL/well Circular wells, V bottom
384 wells 112 µL/well Square wells, flat bottom

API load name appliedbiosystemsmicroamp384 wellplate_40ul biorad_384_wellplate_50ul
corning_384_wellplate_112ul_flat

Well plate adapters
The aluminum plates listed below are thermal adapters for the Opentrons Heater-Shaker GEN1 module. You can use these standalone adapter definitions to load Opentrons verified or custom labware on top of the Heater-Shaker.

Adapter type Opentrons Universal Flat Heater-Shaker Adapter Opentrons 96 PCR Heater-Shaker Adapter Opentrons 96 Deep Well Heater-Shaker Adapter Opentrons 96 Flat Bottom Heater-Shaker Adapter

API load name opentrons_universal_flat_adapter opentrons_96_pcr_adapter opentrons_96_deep_well_adapter opentrons_96_flat_bottom_adapter

You can also load both the adapter and labware with a single definition. Our Labware Library includes several pre-configured thermal adapter and labware combinations that make the Heater-Shaker ready to use right out of the box.

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Note: Don’t use a combined definition if you need to move labware onto or off of the Heater-Shaker during your protocol, either with the gripper or manually. Use a standalone adapter definition instead.

Adapter/labware combination

API load name

Opentrons 96 Deep Well Heater-Shaker Adapter with NEST Deep Well Plate 2 mL

opentrons_96_deep_well_adapternest wellplate_2ml_deep

Opentrons 96 Flat Bottom Heater-Shaker Adapter with NEST 96 Well Plate 200 µL Flat

opentrons_96_flat_bottom_adapternest wellplate_200ul_flat

Opentrons 96 PCR Heater-Shaker Adapter with NEST Well Plate 100 µL

opentrons_96_pcr_adapter_nestwellplate 100ul_pcr_full_skirt

Opentrons Universal Flat Heater-Shaker Adapter with Corning 384 Well Plate 112 µL Flat

opentrons_universal_flat_adapter_corning384 wellplate_112ul_flat

Adapters can be purchased directly from Opentrons at https://shop.opentrons.com.

Well plates and API definitions
The Opentrons Labware Library defines the characteristics of the well plates listed above in separate JSON files. The Flex robot and the Opentrons Python API rely on these JSON definitions to work with labware used by your protocols. For example, when working with the API, the ProtocolContext.load_labware function accepts these labware names as valid parameters in your code. Linked API load names connect to the well plate labware definitions in the Opentrons GitHub repository.

Custom well plate labware
Try using the Opentrons Labware Creator to make a custom labware definition if a well plate you’d like to use isn’t listed here. A custom definition combines all the dimensions, metadata, shapes, volumetric capacity, and other information in a JSON file. The Opentrons Flex reads this information to understand how to work with your custom labware. See the Custom Labware Definitions section for more information.

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5.4 Tips and tip racks
Opentrons Flex tips come in 50 µL, 200 µL, and 1000 µL sizes. These are clear, non-conducting polypropylene tips that are available with or without filters. They’re packaged sterile in racks that hold 96 tips and are free of DNase, RNase, protease, pyrogens, human DNA, endotoxins, and PCR inhibitors. Racks also include lot numbers and expiration dates.
Flex pipette tips work with the Opentrons Flex 50 µL and 1000 µL pipettes in the 1-, 8-, and 96-channel configurations. While you can put any of the Flex tips on the 50 L and 1000 L pipettes, try to match the tip to the pipette’s rated capacity. For example, it might be odd to put a 1000 L tip on the 50 L pipette. For the 1000 L pipette, you could certainly use a 50 L, 200 L, or 1000 L tip.

Tip racks
Unfiltered and filtered tips are bundled into a rack that consists of a reusable base plate, a mid-plate that holds 96 tips, and a lid.

Tip rack by volume 50 µL 200 µL 1000 µL

API load name
Unfiltered: opentrons_flex_96_tiprack_50ul Filtered: opentrons_flex_96_filtertiprack_50ul
Unfiltered: opentrons_flex_96_tiprack_200ul Filtered: opentrons_flex_96_filtertiprack_200ul
Unfiltered: opentrons_flex_96_tiprack_1000ul Filtered: opentrons_flex_96_filtertiprack_1000ul

To help with identification, the tip rack mid-plates are color coded based on tip size:
50 µL: magenta 200 L: yellow 1000 µL: blue

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When ordering or reordering, tips and racks come in two different packaged configurations:
Racks: Consist of separately shrink-wrapped tip racks (base plate, mid-plate with tips, and lid). Racked configurations are best when cleanliness is paramount, to avoid cross-contamination, or when your protocols don’t allow for base plate or component reuse.
Refills: Consist of one complete tip rack (base plate, mid plate with tips, and lid) and individual tip containers. Refill configurations are best when your protocols allow for base plate or component reuse.
Tip­pipette compatibility
Flex pipette tips are designed for the Opentrons Flex pipettes. Flex tips are not backwards compatible with Opentrons OT-2 pipettes, nor can you use OT-2 tips on Flex pipettes.
Other industry-standard tips may work with Flex pipettes, but this is not recommended. To ensure optimum performance, you should only use Opentrons Flex tips with Flex pipettes.
Tip rack adapter
The 96-channel pipette requires an adapter to attach a full rack of tips properly. During the attachment procedure, the pipette moves over the adapter, lowers itself onto the mounting pins, and pulls tips onto the pipettes by lifting the adapter and tip rack.

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Note: Only use the tip rack adapter when picking up a full rack of tips at once. Place tip racks directly on the deck when picking up fewer tips.
Warning: Pinch point hazard. Keep hands away from the tip rack adapter while the pipette is attaching pipette tips.

Adapter type Opentrons Flex 96 Tip Rack Adapter

API load name opentrons_flex_96_tiprack_adapter

The tip rack adapter is compatible with the Opentrons Flex Gripper. You can use the gripper to place fresh tip racks on the adapter or to pick up and move used tip racks into the waste chute.

5.5 Tubes and tube racks

The Opentrons 4-in-1 Tube Rack system works with the Opentrons Flex by default. Using the 4-in-1 tube rack helps reduce your prep work burden because the combinations it provides are automation-ready right out of the box. More information is also available in the Opentrons Labware Library.

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Tube and rack combinations
The Opentrons 4-in-1 tube rack supports a wide variety of tube sizes, singly or in different size (volume) combinations. These include a: 6-tube rack for 50 mL tubes (6 x 50 mL). 10-tube combination rack for four 50 mL tubes and six 15 mL tubes (4 x 50 mL, 6 x 15 mL). 15-tube rack for 15 mL tubes (15 x 15 mL). 24-tube rack for 0.5 mL, 1.5 mL, or 2 mL tubes (24 x 0.5 mL, 1.5 mL, 2 mL).
Note: All tubes are cylindrical with V-shaped (conical) bottoms unless otherwise indicated.

6-tube racks
Tube type 6 Falcon 50 mL 6 NEST 50 mL
10-tube racks
Tube type 4 Falcon 50 mL 6 Falcon 15 mL 4 NEST 50 mL 6 NEST 15 mL

API load name opentrons_6_tuberack_falcon_50ml_conical opentrons_6_tuberack_nest_50ml_conical
API load name opentrons_10_tuberack_falcon_4x50ml_6x15ml_conical opentrons_10_tuberack_nest_4x50ml_6x15ml_conical

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15-tube racks
Tube type 15 Falcon 15 mL 15 NEST 15 mL

API load name opentrons_15_tuberack_falcon_15ml_conical opentrons_15_tuberack_nest_15ml_conical

24-tube racks

Tube type
24 Eppendorf Safe-Lock 1.5 mL 24 Eppendorf Safe-Lock 2 mL, U-shaped bottom 24 generic 2 mL screw cap 24 NEST 0.5 mL screw cap 24 NEST 1.5 mL screw cap 24 NEST 1.5 mL snap cap 24 NEST 2 mL screw cap 24 NEST 2 mL snap cap, U-shaped bottom

API load name opentrons_24_tuberack_eppendorf_1.5ml_safelock_snapcap opentrons_24_tuberack_eppendorf_2ml_safelock_snapcap
opentrons_24_tuberack_generic_2ml_screwcap opentrons_24_tuberack_nest_0.5ml_screwcap opentrons_24_tuberack_nest_1.5ml_screwcap opentrons_24_tuberack_nest_1.5ml_snapcap opentrons_24_tuberack_nest_2ml_screwcap opentrons_24_tuberack_nest_2ml_snapcap

Tube rack API definitions
The Opentrons Labware Library defines the characteristics of the tube racks listed above in separate JSON files. The Flex robot and the Opentrons Python API rely on these JSON definitions to work with labware used by your protocols. For example, when working with the API, the ProtocolContext.load_labware function accepts these labware names as valid parameters in your code. Linked API load names connect to the tube rack labware definitions in the Opentrons GitHub repository.

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Custom tube rack labware
Try creating a custom labware definition using the Opentrons Labware Creator if a tube and rack combination you’d like to use isn’t listed here. A custom definition combines all the dimensions, metadata, shapes, volumetric capacity, and other information in a JSON file. The Opentrons Flex reads this information to understand how to work with your custom labware. See the Custom Labware Definitions section section for more information.
5.6 Aluminum blocks
Aluminum blocks ship with the Temperature Module GEN2 and can be purchased separately as a three-piece set. The set includes a flat bottom plate, a 24-well block, and a 96-well block.
The Opentrons Flex uses aluminum blocks to hold sample tubes and well plates on the Temperature Module or directly on the deck. When used with the Temperature Module, the aluminum blocks can keep your sample tubes, PCR strips, or plates at a constant temperature between 4 °C and 95 °C.
Flat bottom plate
The flat bottom plate for Flex ships with the Temperature Module’s caddy and is compatible with various ANSI/SLAS standard well plates. This flat plate differs from the plate that ships with the Temperature Module itself or the separate three-piece set. It features a wider working surface and chamfered corner clips. These features help improve the performance of the Opentrons Flex Gripper when moving labware onto or off of the plate.
You can tell which flat bottom plate you have because the one for Flex has the words “Opentrons Flex” on its top surface. The one for OT-2 does not.

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24-well aluminum block
The 24-well block is used with individual sample vials. For example, it accepts sample vials that:
Have V-shaped or U-shaped bottoms. Secure contents with snap cap or screw cap closures. Hold liquid in capacities of 0.5 mL, 1.5 mL, and 2 mL.

96-well aluminum block
The 96-well block supports a wide variety of well plate types. For example, it accepts well plates that are:
From major well-plate manufacturers like Bio-Rad and NEST.
Designed with V-shaped bottoms, U-shaped bottoms, or flat bottoms.
Designed with 100 µL or 200 µL wells.
It is also compatible with generic PCR strips.

Standalone adapters
Thermal block Flex flat bottom plate 24-well aluminum block 96-well aluminum block

API load name opentrons_aluminum_flat_bottom_plate See labware combinations below. opentrons_96_well_aluminum_block

Aluminum block labware combinations
The Opentrons Labware Library supports the following block, vial, and well plate combinations, which are also defined in separate JSON labware definition files. The Flex robot and the Opentrons Python API

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rely on these JSON definitions to work with labware used by your protocols. For example, when working with the API, the ProtocolContext.load_labware function accepts these labware names as valid parameters in your code. The tables below list the default block/container combinations and related API load names. Links connect to corresponding JSON definitions in the Opentrons GitHub repository.
Note: All tubes have V-shaped bottoms unless otherwise indicated.

24-well aluminum block labware combinations

24-well block contents Generic 2 mL screw cap NEST 0.5 mL screw cap NEST 1.5 mL screw cap NEST 1.5 mL snap cap NEST 2 mL screw cap NEST 2 mL snap cap, U-shaped bottom

API load name opentrons_24_aluminumblock_generic_2ml_screwcap opentrons_24_aluminumblock_nest_0.5ml_screwcap opentrons_24_aluminumblock_nest_1.5ml_screwcap opentrons_24_aluminumblock_nest_1.5ml_snapcap opentrons_24_aluminumblock_nest_2ml_screwcap opentrons_24_aluminumblock_nest_2ml_snapcap

96-well aluminum block labware combinations

96-well block contents Bio-Rad well plate 200 µL Generic PCR strip 200 µL NEST well plate 100 µL

API load name opentrons_96_aluminumblock_biorad_wellplate_200uL opentrons_96_aluminumblock_generic_pcr_strip_200uL opentrons_96_aluminumblock_nest_wellplate_100uL

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5.7 Labware and the Opentrons Flex Gripper
Although the Opentrons Flex works with all the inventory in the Labware Library, the Opentrons Flex Gripper is compatible with specific labware items only. Currently, the Gripper is optimized for use with the following labware items.

Labware category Deep Well Plates Fully Skirted 96 Well Plates
Tip Racks (unfiltered and filtered tips)

Brands
NEST 96 Deep Well Plate 2 mL
Opentrons Tough 96 Well Plate 200 µL PCR Full Skirt NEST 96 Well Plate 200 µL Flat
Opentrons Flex 96 Tip Rack 50 µL Opentrons Flex 96 Tip Rack 200 µL Opentrons Flex 96 Tip Rack 1000 µL

Note: For best results, use the Flex Gripper only with the labware listed above. The Flex Gripper may work with other ANSI/SLAS automation compliant labware, but this is not recommended.

5.8 Custom labware definitions
As discussed at the beginning of this chapter, custom labware is labware that’s not listed in the Opentrons Labware Library. You can use other common or unique labware items with the Flex by accurately measuring and recording the characteristics of that object and saving that data in a JSON file. When imported into the app, the Flex and the API uses that JSON data to interact with your labware. Opentrons provides tools and services, which we’ll examine below, to help you use the Flex with custom labware.

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Creating custom labware definitions
Opentrons tools and services help put custom labware within your reach. These features accommodate different skill levels and ways of working. Creating your own labware, and using it with the Opentrons Flex, helps make the robot a versatile and powerful addition to your lab.

CUSTOM LABWARE CREATOR
The Custom Labware Creator is a no-code, web-based tool that uses a graphical interface to help you create a labware definition file. Labware Creator produces a JSON labware definition file that you import into the Opentrons App. After that, your custom labware is available to the Flex robot and the Python API.

CUSTOM LABWARE SERVICE
Get in touch with us if the labware you’d like to use isn’t available in the library, if you can’t create your own definitions, or because a custom item includes different shapes, sizes, or other irregularities described below.

Labware you can define in Labware Creator
; Wells and tubes are uniform and identical. ; All rows are evenly spaced
(the space between rows is equal).
; All columns are evenly spaced (the space between columns is equal).
; Fits perfectly in one deck slot.

Labware Opentrons needs to define ; Wells and tube shapes vary. ; Rows are not evenly spaced.
; Columns are not evenly spaced.
; Smaller than one deck slot (requires adapter) or spans multiple deck slots.

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Here are some diagrams that help you visualize the examples described above. Regular All columns are evenly spaced and all rows are evenly spaced. Columns do not need to have the same spacing as rows.
Regular The grid does not have to be in the center of labware.
Irregular Rows are evenly spaced but columns are not evenly spaced.
Irregular Columns/rows are evenly spaced but wells are not identical.
Irregular There is more than one grid.
Our labware team will work to understand your needs and design custom labware definitions for you. See the support articles Requesting a Custom Labware Definition and the Custom Labware Request form for more information. This is a fee-based service.

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PYTHON API
While you cannot create custom labware with our API, you can use custom labware with the available API methods. However, you need to define your custom labware first and import it into the Opentrons App. Once you have added your labware to the Opentrons App, it’s available to the Python API and the robot. See the Custom Labware Definitions section of the Python API documentation for more information. For information about writing protocol scripts with the API, see the Python Protocol API section in the Protocol Development chapter.
JSON labware schema
A JSON file is the blueprint for Opentrons standard and custom labware. This file contains and organizes labware data according to the design specifications set by the default schema.
A schema is a frame

References

• P65Warnings.ca.gov

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