IMPLEN N50-Go Nano Photo Meter User Manual

June 10, 2024
IMPLEN

IMPLEN N50-Go Nano Photo Meter

IMPLEN-N50-Go-Nano-Photo-Meter

Product Information

The product is a spectrophotometer with a software version of 4.6.16350 and a user manual version of 4.6.2. It is manufactured by Implen, Inc. and comes with telephone support available for customers in North and South America. The product meets various safety and environmental standards, including the Low Voltage
Equipment Safety Directive, Electromagnetic compatibility (EMC) directive, Restrictions on the use of certain Hazardous Substances in Electrical and Electronic Equipment (ROHS), and EC Directive on Waste Electrical and Electronic Equipment (WEEE).

Product Usage Instructions

  1. Accessories: The product comes with standard and optional accessories. Refer to the user manual for a detailed list of accessories.
  2. Getting Started:
    • Spectrophotometer Installation: Follow the instructions in the user manual for proper installation of the spectrophotometer.
    • Safety Information: Read the safety information section in the user manual before using the product.
    • Unpacking and Positioning: Follow the instructions in the user manual for proper unpacking and positioning of the product.
  3. Icons: Refer to the icons section in the user manual to understand the meaning of different icons displayed on the product screen.
  4. Buttons: Refer to the buttons section in the user manual to understand the function of different buttons on the product.
  5. Measurement Screens: Refer to the measurement screens section in the user manual to understand how to read and interpret different measurement screens displayed on the product.
  6. Methods: Refer to the methods section in the user manual to understand how to use different methods for measuring samples using the product.

NanoPhotometer® N50-Go/C40-Go
User Manual
Version 4.6.2 Software Version 4.6.16350

NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2 The end user of the NanoPhotometer® Go product (“End User”) hereby takes full responsibility for safe storage and backup of all files and/or data that may be created, saved on or transferred from the device. End User acknowledges that it is possible that data and/or files may be lost or damaged, and further acknowledges and agrees that it has sole responsibility to maintain all appropriate backup of files and data. By using the NanoPhotometer® Go device, End User hereby agrees to these terms, and agrees that Implen shall not be held liable for any loss, deletion or damage of any data or files for any reason, including any damages attributable thereto.
Telephone support is available using one of the following phone numbers from your geographic region: North and South America
Phone: +1 818 748 6400 Fax: +1 818 449-0416 Email: info@implen-go.com Website: www.implen-go.com Implen, Inc. Unit 104 31194 La Baya Drive Westlake Village, CA 91362 USA
Windows and Excel are trademarks of Microsoft Corporation Redmond, WA macOS is a trademark of Apple Inc. Cupertino, CA
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2

Declaration of conformity for the NanoPhotometer® (C40-Go/N50Go)

This is to certify that the Implen NanoPhotometer® Go conforms to the requirements of the following directives:

2014/35/EU

Low Voltage Equipment Safety Directive

2014/30/EU

Electromagnetic compatibility (EMC) directive

IEC 60529

Protection class IP20

2011/65/EU

Restrictions on the use of certain Hazardous Substances in Electrical and Electronic Equipment (ROHS)

2012/19/EU

EC Directive on Waste Electrical and Electronic Equipment (WEEE) 2003/108/EC & 2008/34/EC. By ensuring this product is disposed of correctly, you will help prevent potential negative consequences for the environment and human health, which could otherwise be caused by inappropriate waste handling of this product.

FCC 47 CFR Part15 §15.107 and §15.109

EN 301 489-1 V1.9.2

Radio and ancillary equipment for portable use (portable equipment); EUT Operating frequency range: 2.4 ­ 2.4835 GHz

EN 300 328 V1.8.1

Electromagnetic compatibility and Radio spectrum Matters (ERM); Wideband transmission systems; Data transmission equipment operating in the 2,4 GHz ISM band and using wide band modulation techniques; Harmonized EN covering the essential requirements of article 3.2 of the R&TTE Directive

EN 301 489-17 V2.2.1

Electromagnetic compatibility and Radio Spectrum Matters (ERM)

Standards to which conformity is declared, where relevant, are as follows:

IEC/EN 61010-1:2012 EN61326-1:2013

Safety requirements for electrical equipment for measurement, control, and laboratory use. General requirements.
Electromagnetic compatibility- generic emission standard electrical equipment for measurement, control, and laboratory use.

Signed:

Dr. Thomas Sahiri Managing Director Implen GmbH

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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2
NANOPHOTOMETER® N50-GO
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2
NANOPHOTOMETER® C40-GO
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2
INSTRUMENT REAR PANEL
To boot/shut down the NanoPhotometer® Go push shortly (< 1 second) on the power on/off button at the back of the NanoPhotometer® Go. Note: A long push (> 3 seconds) initiate a hard reset. Only activate a hard reset of the NanoPhotometer® Go when necessary. To avoid unnecessary hard reboots, it is recommended to power down the unit from the onboard touch screen by pushing on the power button ( ) in the bottom left corner of the home screen.
INSTRUMENT BOTTOM VIEW
Model name, device serial number and FCC ID are located on the identification plate on the bottom of the instrument.
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2

ACCESSORIES

STANDARD ACCESSORIES
Connecting Cable
USB connecting cable to connect the NanoPhotometer® Go to a computer to control the NanoPhotometer® Go via computer (please see page 21 Software Installation).
NanoPhotometer® Go Power Adapter
Power adapter for the NanoPhotometer® Go. Note: Use only the power adapter supplied with your instrument or a replacement part from the manufacturer or your supplier.
Dust Cover
OPTIONAL ACCESSORIES
Didymium Glass Filter (C40-Go) The certified didymium glass filter can be used to verify the wavelength and photometric accuracy of the NanoPhotometer® C40-Go cuvette applications.
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2
Standard Solution (N50-Go)
The NanoPhotometer® Go is recalibration free therefore it is not necessary to review the photometric accuracy on a regular basis. If the SOPs of a laboratory require a routine control of the photometric accuracy standard solutions can be used.

Note: Specifications of the standard solution are guaranteed for at least one year. Please see expiration date. Once a vail is opened it can be used for 30 minutes. Note: Please read the Material Safety Data Sheet carefully prior to using this product.

DiluCellTM (C40-Go)
The DiluCellTM is a disposable cuvette with shortened path length for virtual dilution of cuvette-based samples. Due to the reduced path length the DiluCellTM provides an automatic dilution without the need of a physical dilution of higher concentration samples. There are two different sizes of DiluCells available DC 10 and DC 20 which allow an automatic 1/10 and 1/20 dilution of the sample respectively. Bypassing manual sample dilutions reduces dilution errors and cross-contamination making DiluCellTM ideal for GLP. Combined with small sample volume requirements and bubble free filling, the DiluCellTM allows for convenient spectrophotometric analysis from 340 – 950 nm.

Barcode Reader
It is possible to import sample names from 1D and 2D barcodes. Connect a compatible barcode reader to the USB port of the NanoPhotometer® Go and push on the sample name input window. After scanning a barcode the sample name will be shown in the input window. The imported name can then be edited or replaced completely.

Barcode readers that have been tested and verified compatible:

1D:

Honeywell Voyager 95X0 Single-Line Laser Scanner

Datalogic Touch65

1D & 2D: AGPtEK SC36

Honeywell Xenon 1900g

DYMO Label Printer It is possible to connect a DYMO Label printer to the NanoPhotometer® Go for direct printing on standard or cryo labels. Recommended and tested printers are the DYMO LabelWriter 4XL/5XL (label size 10.3 x 15.8 cm) and the DYMO LabelWriter 450/550 (5.4 x 10 cm).
Cryo labels can be printed with DYMO Label printer 4XL and 450 using the following label format: 26 x 12.7 mm and 9.5 mm circle (landscape).

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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2 Note: Cryo label paper is not available/compatible for the DYMO Label printer 5XL and 550. Note: After starting the NanoPhotometer® Go, plug in the DYMO printer. The home screen will be shown. Wait at least for 30 seconds for driver installation. Cryo labels (26 x 12.7 mm and 9.5 mm circle):
Note: Orientation of cryo labels on carrier foil needs to be landscape.

Printout for DYMO LabelWriter 4XL/5XL

Printout for DYMO LabelWriter 450/550

Note: The printouts are optimized for the label size of the DYMO LabelWriter 4XL. All other DYMO printers can be used. However, the font size will be zoomed to utilized paper size.

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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2

HP Printer
Printing from the NanoPhotometer® Go is possible via USB (HP printers) and through network connection. Network printing should be possible with AirPrint / IPP compatible printers supporting PDF format. For network printer settings see page 96 Network Printer.

Note: IPP version 2.2 is required and some printer configuration settings might need to be changed in order to allow communication with the NanoPhotometer® Go.

The following HP printers have been tested and deemed compatible to print via USB

connection:

HP LaserJet 3030

HP DeskJet 2543

HP LaserJet m1522nf MFP

HP DeskJet 1110

HP LaserJet 400 color M451nw

Further HP printers are available upon request.

Note: After starting the NanoPhotometer® Go, plug in the HP printer. The home screen will be shown. Wait at least for 30 seconds for driver installation.

CONNECTIVITY

USB A
There is a USB A port on the front as well as the rear panel of the NanoPhotometer® Go which is compatible with standard portable USB 2.0 storage devices (back) and USB 3.0 (front) for direct data transfer in a variety of formats including Excel. It is also possible to connect a mouse, keyboard, barcode reader, DYMO printer or HP printer directly to the NanoPhotometer® Go.
Note: We recommend using FAT/FAT32 formatted 2.0 USB flash drives. The USB flash drive size is due to the standard formatting currently limited to 32 GB. Encrypted USB flash drives are not compatible with the NanoPhotometer® Go. Note: Cordless Bluetooth mice are not supported. Use wired mice only. Note: Connect mouse and keyboard before starting the NanoPhotometer® Go.
USB B
There is a USB B port located on the rear panel of the instrument which is compatible with the USB cable provided to connect the NanoPhotometer® Go to a computer. This USB connection can be used to control the NanoPhotometer® Go via computer.
LAN
There is an Ethernet (LAN) connection port on the rear panel of the instrument which enables the NanoPhotometer® Go to connect with the local network. This Ethernet connection can be used for data transfer from the NanoPhotometer® Go to a local network, to control the NanoPhotometer® Go via a control device and network printing. Data transfer is possible by saving on a defined network folder (see page 95 Network Folder) or on the NanoPhotometer® Go file server (see page 38 Data Transfer via File Server).

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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2 Note: Plug in the LAN cable before starting the NanoPhotometer® Go. Note: The maximum LAN cable length is 10 meters. Bit rate is 1 Gbit/s
WiFi The NanoPhotometer® Go is equipped with WiFi, which can be used as a WiFi network or as a WiFi Hotspot. The WiFi network allows same functionality as the Ethernet connection including direct printing via AirPrint / IPP compatible printers supporting PDF format. Note: IPP version 2.2 is required and some printer configuration settings might need to be changed in order to allow communication with the NanoPhotometer® Go. The WiFi Hotspot provides the option to control the NanoPhotometer® Go by other WiFi devices like computer, tablets or smartphones. WiFi Hotspot connection details: SSID: NanoPhotometer® Go serial number Password: Implenuser Note: Due to the limitations of some handheld devices saving to a wireless device is limited to 40 measurements per dataset. Larger datasets can be saved on the NanoPhotometer® Go itself.
HDMI There is an HDMI port located on the rear panel of the NanoPhotometer® Go which is compatible with HDMI 1.4 cables (or better) to connect the NanoPhotometer® Go to HDMI compatible monitors. Note: The maximum HDMI cable length is 5 meters.
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2
NANOPHOTOMETER® GO SPECIFICATIONS

NanoVolume Performance ­ N50-Go

Detection Range dsDNA Detection Range BSA Sample Volume Photometric Range (10 mm equivalent) Path length Dilution Factor

5 ­ 7,500 ng/µl 0.15 ­ 217 mg/ml 0.3 ­ 2 µl 0.1 ­ 150 A
0.67 and 0.07 mm 15 and 140

Cuvette Performance ­ C40-Go

Detection Range dsDNA

0.1 – 130 ng/µl

Detection Range BSA

0.003 – 3.7 mg/ml

Photometric Range

0 – 2.6 A

Center Height (Z-Height)

8.5 mm

Cell Types

outside dimension 12.5 x 12.5 mm

Heating

37°C ± 0.5°C

Optical Specifications Wavelength Scan Range Measure Time For Full Scan Range Wavelength Reproducibility Wavelength Accuracy Bandwidth Absorbance Reproducibility
Absorbance Accuracy Stray Light Optical Arrangement

C40-Go: 200 ­ 900 nm N50-Go: 200 ­ 650 nm 2.5 ­ 4.0 seconds
C40-Go: ± 0.2 nm N50-Go: ± 1nm C40-Go: ± 0.75 nm N50-Go: ± 1.5 nm C40-Go: < 1.5 nm N50-Go: < 3 nm C40-Go (cuvette): < 0.002 A @ 0 – 0.3 A @ 280 nm
CV < 1% @ 0.3 ­ 2.0 A @ 280 nm N50-Go (Lid 15): < 0.004 A (0.67 mm path) @ 280 nm
CV < 1% @ 0,3 – 1,5 A @ 280 nm < 1.75 % @ 0.7 A @ 280 nm of the reading C40-Go: < 0.5% @ 240 nm using NaI N50-Go: < 2% @ 240 nm using NaI 1 x 4096 CMOS Array

Lamp Lifetime

Xenon flash lamp 109 flashes, up to 10 years

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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2

Processing Power and Compatibility

Operating System

Linux based OS

Onboard Processor

Intel Celeron dual core 2.4 GHz

Internal Data Storage

C40-Go, N50-Go: 32 GB

Software Compatibility

Windows 8, 10 (32 & 64 bit), OS X (Intel x86 und Apple M1), iOS and Android OS

General Specifications Main Body Size Weight Operating Voltage Display Certifications In- and Output Ports Security

200 mm x 200 mm x 120 mm 3.8 ­ 5.2 kg depending on configuration 90 ­ 250 V ± 10%, 50/60 Hz, 90 W, 18/19 VDC 1024 x 600 pixels; touchscreen glove compatible CE, IEC 61010-1:2012 and EN 61326-1:2013 2x USB A, USB B, HDMI, Ethernet, WiFi Slot for Kensington lock

Features and specifications are subject to change without notice. US Patents 20080204755 and 20080106742 Windows is a trademark of Microsoft. Mac OS & iOS are trademarks of Apple, Inc. Android OS is a trademark of Google. Linux is a trademark of Linus Torvalds.

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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2
2. GETTING STARTED
SPECTROPHOTOMETER INSTALLATION
SAFETY INFORMATION
Before commencing installation, please take time to familiarize yourself with warning labels and symbols on your instrument and their meaning. These are to inform you where potential danger exists or particular caution is required. Improper use may cause personal injuries or damage to the instrument. The instrument must only be operated by appropriately trained and experienced personnel. Please read the complete user manual prior to use.
direct current Overvoltage catetory: Class II Maximum operating altitude: < 2000 m Polution degree: 2
Do not open the instrument as this can expose the operator to electrical power, UV light, and delicate fiber optics or damage the instrument.
Do not use damaged power cords, accessories, and other peripherals with your NanoPhotometer® Go. Use only the delivered and specified power supply.
Do not expose the NanoPhotometer® Go to strong magnetic, electrical fields, water, chemicals or any type of liquid as heavy rain or moisture.
Do not put the instrument into fire, as it may swell or explode. Do not store at or use near any type of heat source, especially temperatures above 60°C or in an explosive atmosphere.
Do not leave your NanoPhotometer® Go on your lap or near any part of your body to prevent discomfort or injury from heat exposure.
Do not place objects on top of the NanoPhotometer® Go.
Biological samples may contain or have the potential to transmit infectious diseases. Be aware of the health hazard presented by such samples and wear appropriate protective equipment. Handle such samples with the greatest of care and according to applicable regulatory and organization requirements before working with such potential infectious materials. Note: Do not spill any biological samples on instrument components. If spill occurs, disinfect the instrument immediately following your laboratory protocols and the cleaning instruction of the instrument (see page 107 Maintenance).
The symbol on the product, or on the documents accompanying the product, indicates that this appliance may not be treated as household waste. Instead it shall be handed over to the applicable collection point for the recycling of electrical and electronic equipment. Disposal must be carried out in accordance with local environmental regulations for waste disposal.
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2
UNPACKING AND POSITIONING
Check the contents of the package against the delivery note. If any shortages are discovered, inform your supplier immediately. Inspect the instrument for any signs of damage caused in transit. If any damage is discovered, inform your supplier immediately. Ensure your proposed installation site conforms to the environmental conditions for safe operation: indoor use or dry environment. Note: Do not expose your NanoPhotometer® Go near liquids, chemicals, rain, moisture or dusty environments. Working temperature range 10 – 40°C; If the cuvette heating is used the range is 10 – 27°C. Storage temperature range is 0 – 40°C. Do not store the instrument below or above this temperature. If the instrument is subjected to extreme temperature changes, it may be necessary to allow the instrument to equilibrate. Turn the instrument off and then on again once thermal equilibrium has been established (~2 – 3 hours). Maximum relative humidity (non-condensing) of 80% up to 31°C decreasing linearly to 50% at 40°C. The instrument must be placed on a stable, level surface that can support 4-5 kg. Ensure that air can circulate freely around the instrument. Confirm while powered on that no materials reduce air circulation. Avoid direct sunlight as it may bleach parts of the instrument and can cause damage to plastic parts. The equipment should be positioned such that in the event of an emergency the main plug can be easily located and removed. Always carry the instrument by holding the main corpus of the instrument and not e.g. on the optional attached display or NanoVolume pedestal. The equipment must be connected to power with the 90W power supply/cord supplied by Implen. The power outlet must have a protective conductor (earth/ground). It can be used on 90-250 V ± 10%, 50-60 Hz power supply system. Please read the complete user manual before first use. Turn the instrument on using the power button on the rear panel after it has been plugged in. The instrument will perform a series of self-diagnostic checks. Please contact original supplier immediately if technical or sample handling difficulties are experienced. Note: If this equipment is used in a manner not specified or in environmental condition not suitable for safe operation, the protection provided by the equipment may be impaired and the instrument warranty voided.
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2
SOFTWARE INSTALLATION
NPOS OVERVIEW
NPOS is a Linux-based operating system designed for the NanoPhotometer® Go.
NPOS can store data either to a common directory or be configured to save to independent directories according to file format and/or instrument.
NPOS can save data in an Implen IDS format, PDF or as an Excel format file.
Note: PDF and Excel files cannot be opened on the NanoPhotometer® Go. Files need to be transferred to a computer or device where Excel or a PDF reader is available. Note: Please do not connect the instrument to a computer until the NanoPhotometer® Go NPOS software is installed on the computer.
REQUIREMENTS AND COMPATIBILITY
The NPOS user interface is designed that all features can be operated by using a touchscreen. If the software is installed on a computer without touchscreen, the user interface can be operated by using keyboard and mouse. Before starting the installation process, ensure that the software of the control device is compatible.
Compatible Control Devices
Computer: PC: Windows 8 / Windows 10 (32 & 64 bit) Mac: macOS Catalina / Big Sur (Intel x86 und Apple M1) Tablets (minimum requirements): iPad: iOS 13 Android (quad core 1.2 GHz with 1 GB RAM): Android version 10 Smartphones (minimum requirements): iPhone: iOS 13 Android (quad core 1.2 GHz with 1 GB RAM): Android version 10
Windows is a trademark of Microsoft. Mac OS & iOS are trademarks of Apple. Android OS is a trademark of Google. Linux is a trademark of Linus Torvalds.
Note: There are two user interfaces of the software available; one for built- in touchscreen, computer and tablets and one for smartphones.
INSTALLING SOFTWARE ON COMPUTER
The NanoPhotometer® Go software can be installed on compatible Windows and Mac computer systems. Various operating systems and computer hardware may cause the setup procedure to differ from that described here. This process is given as guidance only; it may need adaptation for other systems.
Note: Do not connect the NanoPhotometer® Go to the PC/Mac before NPOS installation. Note: If a previous version of the NPOS software is already installed on the computer, remove the USB cable and uninstall the NPOS software before installing the new software
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2 version. Note: The Windows and Mac installation files are located on the Implen USB flash drive which is included with the NanoPhotometer® Go at time of delivery. The files are available for free download at any time in the download area of the Implen website (www.implen.de/downloads).
NPOS installation for single user on Windows computer 1. Check the installed version of the NanoPhotometer® Go firmware
(Preferences/About) and update it to the latest version if necessary, before starting the installation/update of the NPOS software on your computer. 2. Start the NPOS installation file and follow the installation routine for single user installation. Installation file can be found on the Implen USB flash drive which is included with the NanoPhotometer® Go at time of delivery or can be downloaded from the Implen webpage: www.implen.de/downloads. Full administration rights are required for the installation. If you have insufficient privileges, installation may fail. If in doubt consult your computer administrator. 3. For multi user installation (only necessary for Windows installation) choose on the License Agreement dialog the option “Advanced” and on the following dialog “Install for all users of this machine”
4. Start the NPOS software and select the desired connection. For a connection via USB cable, connect the NanoPhotometer® Go to the PC using the USB cable supplied. For a connection via WiFi hotspot, ensure a stable WiFi connection between the PC and NanoPhotometer® Go WiFi HotSpot (SSID: serial number, password: Implenuser). For a network connection, connect the NanoPhotometer® Go to the local network via an Ethernet cable or WiFi network (see page 92 Network).
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2
Note: If there is Avira installed on your computer, it is recommended to switch off the browser safety. This may interfere with the NPOS running on your computer.
INSTALLING NANOPHOTOMETER® GO APP ON TABLET OR SMARTPHONE
The NanoPhotometer® Go App can be installed as an application on tablets and smartphones with compatible Android and iOS operating systems (see page 21 Requirements and Compatibility). The NanoPhotometer® Go App is available for free download in the app store (Apple Store and Google Play Store).
1. Download and install the NanoPhotometer® Go App from the app store 2. Connect the tablet or smartphone via WiFi HotSpot (SSID: Serial number,
password: Implenuser) or WLAN network to the NanoPhotometer® Go. 3. Open the NanoPhotometer® Go App and choose the connection type:
4. When connected the NanoPhotometer® Go will recognize the tablet/smartphone as a remote control device and measurements can be initiated from the tablet or smartphone.
5. Results will be shown on the tablet or smartphone once measurements have been taken.
Note: In order to install the NanoPhotometer® Go App on a tablet or smartphone, the device must have an established internet connection to access the app store for app download. Note: The version of the app and the software of the NanoPhotometer® Go should be the same. Different versions may have not full functionality.
FIRST STEPS AND CONFIGURATION WIZARD
When starting the Implen NPOS the first time an Implen configuration wizard is shown. Please accept the End User License Agreement (EULA) and select the country in which the NanoPhotometer® Go is used and confirm.
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2
PRINTER INSTALLATION
For printers connected via USB connection: 1. Switch NanoPhotometer® Go on / home screen 2. Connect DYMO/HP printer via USB cable 3. DYMO/HP printer is ready to use after 30 seconds
Note: Make sure that the home screen is shown when connecting a printer. If the USB connection between the printer and the NanoPhotometer® Go is established while a method is open, the printer function may fail. Always return to the home screen before connecting a printer. Note: Check printer compatibility (page 15 HP Printer) For network printers:
1. Assure either LAN or WiFi network connection 2. Set printer IP in preferences (page 96 Network Printer) 3. Printer is available in methods for printing Note: Check printer compatibility (page 15 HP Printer)
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2
3. NANOPHOTOMETER® GO BASICS
The NanoPhotometer® Go product line offers a solution for NanoVolume (N50-Go) and standard cuvette (C40-Go) applications. NanoVolume applications start with a minimum sample volume of 0.3 l. Standard cuvette applications can be performed with 10 mm, 5 mm, 2 mm, 1 mm and 0.5 mm path length quartz, glass, or plastic cuvettes with a center height of 8.5 mm.
APPLICATIONS OVERVIEW
The NanoPhotometer® N50-Go comes with two pre-programmed applications, Nucleic Acids and Protein UV. To select a method, tap the corresponding icon and the method opens immediately.
There is an upgrade option for the N50-Go available. The upgraded N50-Go is a N50 Touch with access to all applications available. Homescreen C40-Go:
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2

The following methods are available for the upgraded N50-Go and the C40-Go. *not available for N50-Go

Method Icons

Description

Nucleic Acids

Concentration, purity, and dye incorporation for DNA, RNA, Oligo, and other nucleic acids

Protein UV

Protein UV determination at 280 nm (or in a range of 200 330 nm), purity and dye incorporation

Kinetics*

Time vs. Absorbance readings

Protein Assays*

BCA (562 nm), Bradford (595 nm), Lowry (750 nm), and Biuret (546 nm) Assays

OD600*

Measures cell density at 600 nm (or in a range of 200 ­ 900 nm)

Stored Results Archive of stored results

Stored Methods Collection of stored custom methods

More Apps*

Additional applications found on a secondary method screen

Wavelength Wavescan

Define one or multiple wavelength between 200 ­ 900 nm (N50-Go: 200 ­ 650 nm) for absorbance measurements
Define desired full scan range anywhere between 200 – 900 nm (N50-Go: 200 – 650 nm)

Concentration*

Define extinction coefficient for automatic concentration calculations

Absorbance/ Ratio*

Define two wavelengths absorbance/ratio calculation

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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2

Standard Curve* Create a standard curve at a defined wavelength

Custom Apps*

Optional custom applications for personalized methods tailored to individual spectroscopy needs

ICONS

Icon Name

Action

WiFi Network WiFi network active; status of WiFi connection

WiFi Hotspot WiFi hotspot active

Assistance

Opens the assistance page

Preferences Opens the preferences page

Home Screen Store Method Save Data

Returns to home screen with application icons for method selection.
Opens a dialog pop up with the possibility to store the actual method parameter to a custom method
Opens a save dialog pop up

Leave Method Returns to the previous application selection

Back

Returns to the previous page (smartphone only)

Next/Confirm Confirms parameter and opens the next screen (smartphone only)

Print Data

Opens a print dialog pop up (only shown when a printer is available)

Delete Data Opens a delete dialog pop up

Parameter

Opens parameter window

Results

Opens result window

Graph

Opens graph result window

Table

Open/shows results in table format

Add Folder Manage Data Delete

Adds a new folder to the directory
Opens a dialog pop up with several action options including delete, rename or import folders/files/data as well as copying or move folders/files/data to defined directories
Deletes added functions in parameter; empties input windows

Full Scale Cancel

Restores graph to original size without zoom
Returns to the previous screen/closes window without implementing any changes

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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2

BUTTONS

When opening a method and starting a sample measurement, a blank measurement is required. For the blank measurement, either water or sample buffer can be used to give the NanoPhotometer® Go a reference of what zero should be. It is recommended to re-apply the blank solution and measure it as a sample to ensure the graph of the blank spectra is a flat line.
To initiate the spectral scan of the sample push the sample button. The data will be temporarily stored until the method is exited; at this time the user needs to define if the samples should be saved.
The auto sample button is disabled by default. When switched on sample measurements are started automatically as soon as the lid arm is closed. Auto sample function is only available for sample and not for blank measurements.

MEASUREMENT SCREENS

METHODS
Side Tab Bar
On the left side of the measurement screen there is a vertical tab bar that contains four tabs including: parameters, results, graph, and table. The different tabs allow the user to organize the measurement screen. It is possible to show or hide the different areas on the screen. Default screen for computer shows all areas, for the built-in screen and the tablet version the table is hidden.

Parameter
Results
Graph
Table
Note: There is no tab bar available for smartphone versions. The parameter, results and graph screens are shown full screen. Parameters need to be confirmed ( ) to get to the measurement screen. It is possible to toggle between the results and graph area by swiping left and right. There is no table area available for smartphones.
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Parameter area In the parameter area it is possible to define all necessary parameters for a measurement as well as turn on cuvette mode and initiate cuvette heating. The standard measurement screen shows the parameter area open by default. The parameter area is automatically hidden when starting either a blank or sample measurement by pushing on the Blank or Sample button. It is also possible to hide the parameter area by tapping the parameter tab in the vertical side tab bar.
Results area The results area shows the method specific results of the grey highlighted measurement in the table including concentration, absorbances, and relevant ratios. It is also possible to change the units of the calculated concentrations in the results area with a dropdown selection menu.
Table area The table area collects the results of all samples in an active method. The first table column shows a tick box. Selecting the samples with the tick box the graphs are overlaid in the graph area. With the header tick box it is possible to select/unselect all samples (maximum number of sample selection is 30). The second column of the table indicates whether the measurement is saved ( ) or not saved (blank field).
With the edit button the sample name of a single sample can be edited.
1. Select one sample in the table (selected sample will be highlighted in grey)
2. Push on Edit button
3. Change sample name
4. Confirm with the “Confirm” button Note: It is not possible to edit sample names of opened IDS files.
Graph area The graph area shows a chart with the graph of the actual measurement or the selected line(s) in the table (tick box selection). There is an overlay toggle switch on the left bottom of the graph area. If the overlay option is enabled the graphs of the measurements will be automatically overlaid. To change the overlaid graphs use the tick boxes in the table.
Note: It is only possible to overlay up to 30 graphs in a chart. If more than 30 data are selected a message will appear that says “More than 30 samples have been selected. Only 30 will be shown in graph.” Note: The overlay button is not available on the NanoPhotometer® Go touch screen and smartphones, only on tablets and computer versions.
It is possible to zoom in and out any area of the diagram (x- and y-axis). Undo the zoom by pushing the full scale icon ( ).
Note: Maximum zoom is 20 nm for the x-axis and 0.01A for the y-axis.
As legend option the sample name in the table is colored in the same color as the graph in the chart. Pushing on the graph opens a pop up which shows the sample name, wavelength and
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2 absorbance of the selected wavelength. It is possible to display the results of other graphs by changing the sample name with the dropdown option.
DATA PROCESSING DIALOGS
PRINT
Selecting the print icon ( ) opens a full screen dialog window with various print options. The print icon is only shown if a printer is available. The print command is sent primarily to DYMO or HP printers if they are directly connected to the NanoPhotometer® Go via USB cable. If no USB printer is available, the print command is sent to the defined network printer, if configured. Network printers can be configured in the preferences by entering the printer IP (see page 96 Network Printer). All ticked samples are printed. Note: If a printer is directly connected to the NanoPhotometer® via USB, this printer will have the highest priority and will be used by default when selecting Print on NanoPhotometer®. In order to print utilizing a printer on the network, please disconnect the connected USB printer.
Note: The print icon is only shown, if a printer is available. Note: It is only possible to operate one printer at a time. Do not connect more than one printer to the NanoPhotometer® Go. Note: The print option is not available in smartphone apps.
Auto Print If the auto print function is enabled, each measurement will be printed directly after the measurement. Auto print function is available for DYMO printer, HP printer connected via USB cable and network printer.
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2 Note: Default setting for auto print is off. If enabled in one method it is set default on for all methods and needs to be switched off if not required. Note: The auto print function is not available for printing via control device (local computer printer).
Cryo Label Print To print on cryo labels connect a DYMO printer (4XL or 450) to the NanoPhotometer® Go and insert the specified cryo label paper (26 x 12.7 mm and 9.5 mm circle / landscape mode). Note: Cryo label paper is not available/compatible for the DYMO Label printer 5XL and 550.
SAVE
Selecting the save data icon ( ) opens a full screen overlay dialog window with various save options.
By default all samples are ticked in the first column of the table and will be saved. It is possible to select samples for saving by using the tick boxes. The header tick box selects/deselects all samples. Note: In the smartphone app there are always all measurements saved, no selection possible.
Save as Type With the Save as Type option it is possible to specify the file type for saving. File type options include Excel, PDF and Implen Document Source (IDS). It is possible to save different file formats simultaneously. Note: IDS files cannot be saved on control devices. IDS files can only be saved in the NanoPhotometer® storage, defined network folder or on a USB flash drive. It is not possible to save IDS files from opened data. In these cases the IDS tick box is greyed out. Note: PDF and Excel files cannot be opened on the NanoPhotometer® Go. The files need to
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be transferred to a computer or device where Excel or PDF reader is available. Note: All data saved on the NanoPhotometer® Go are stored on an internal micro SD card. It is recommended to back up the data regularly to a hard drive of a computer or network. In the rare case that the micro SD card crashes data loss cannot be ruled out.
Implen Document Source File
The Implen Document Source (IDS) file is a specific file format, which can only be opened with the NPOS software. It is a hardcopy file which cannot be changed. This file type contains all measurement information including raw data, results, values, and parameters.
Note: The saved files contain only the selected samples at the time the file is saved.
Excel File
Measurement data can be saved as Excel file. This file type contains all measurement information including raw data, results, and parameters.
Note: The saved files contain only the selected samples at the time the file is saved.
PDF File
Measurement data can be saved as PDF file. This file type contains all measurement information including raw data, results, and parameters. It is possible to configure the table columns for PDFs and printouts in preferences (page 97 Report Configuration).
Note: The saved files contain only the selected samples at the time the file is saved.
File Name Enter the name for the file. Allowed characters are: A…Z a…z 0…9 , . – ( ) @ ! = _ ~ ; [ ] { } `
Note: Blank character is not allowed.
Storage Shows folder directory to select the save location. Options include: NanoPhotometer®, USB flash drive (if connected), Network folder (if defined) and Control Device. If Control Device is selected the data will be transferred to the control device that is currently connected with the NanoPhotometer® Go.
Note: It is not possible to save IDS files to a control device like computer (PC/Mac), tablets or smartphones.
Auto Save To prevent data loss, all measurements are automatically stored as IDS file on the internal memory of the NanoPhotometer® Go. These backup copies can be found in the Autosave folder of the NanoPhotometer® (Stored Results/NanoPhotometer/Autosave) for up to ten days. Files contain the base name Backup, the method name, and a time/date stamp. After ten days the autosave files are automatically moved to an autosave archive folder. The autosave archive folder can only be accessed via NanoPhotometer® Go file server (see
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page 38 Data Transfer via File Server). Data in the autosave archive folder are not automatically deleted.

The content of the Autosave Archive folder can be deleted via the action button Results:

in Stored

DELETE
Selecting the delete icon ( ) opens a full screen overlay dialog window. All data which are selected (tick) in the first table column will be deleted. The header tick box selects/deselects all samples. Initiate the deletion with the delete button. Confirm the deletion in the following warning message: “Do you want to delete all/selected files?” select cancel (x) will return to the delete menu screen or confirm with delete to delete the selected data. Note: The delete function is not available for the software version designed for smartphones.
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STORE METHODS
With the favorite icon it is possible in each method to save the parameter settings for easy access of custom defined methods. Select the desired parameter settings and open the Stored Method dialog by pushing o the favorite icon .
Enter a method name and select a save location in the folder. Options include: NanoPhotometer®, USB flash drive (if connected) and Network folder (if defined). Push the store button to save the method. Stored Methods can be opened on the homescreen by opening Stored Methods (see page 87 Stored Methods).
BASIC OPERATION
The NanoPhotometer® Go product line offers a solution for NanoVolume (N50-Go) and standard cuvette (C40-Go) applications. NanoVolume applications start with a minimum sample volume of 0.3 l. Standard cuvette applications can be performed with 10 mm, 5 mm, 2 mm, 1 mm and 0.5 mm path length quartz, glass, or plastic cuvettes with a center height of 8.5 mm.
NANOVOLUME MEASUREMENT BASICS (N50-GO)
1. Select a method depending on your sample and set the parameters for the measurement.
2. Ensure that the sample window on pedestal and the mirror in the lid arm are clean. 3. Raise the lid arm and pipette the appropriate amount of blank solution onto the
illuminated sample window on pedestal. The Illumination turns automatically off when the lid arm is lowered.
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Note: Do not overfill the well. Note: The low energy red light (LED) illumination can be switched of in Preferences 4. Lower the lid arm and initiate a blank measurement with the blank button 5. Clean the measurement window and mirror on the lid arm with a slightly wet lint-free tissue. Use water, 70% ethanol or isopropanol if needed.
Note: Make sure that the metal contact face (around the measurement window and the mirror) is clean. Note: Do not use aggressive solvents such as strong acids or bases or organic solvents at any time (see page 38 Solvent Compatibility (N50-Go). If unsure please contact support@implen-go.com for detailed information about your specific reagent/solvent. 6. It is possible to enter a sample name for each sample in the input window “enter sample name”. Note: Allowed characters are: A…Z a…z 0…9 , . – ( ) @ ! = _ ~ ; [ ] { } ` blank character 7. Raise the lid arm and pipette the appropriate amount of sample solution onto the illuminated sample window. Upon completion of measurement raise lid arm, clean the surfaces and apply the next sample. Note: Parameter setting Volume 1 – 2 µl adjusts the path length automatically. The parameter setting Volume 0.3 µl measures only the 0.07 mm path length for higher concentrations (dsDNA > 420 ng/µl / BSA > 12.6 mg/ml). Note: The sample window on pedestal must be clean and residual fluff from any cleaning wipe must be removed for optimum performance.
CUVETTE MEASUREMENT BASICS (C40-GO)
The NanoPhotometer® Go is compatible with standard cuvettes having an 8.5 mm center height. The light path is indicated with two white arrows.
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1. Select a method depending on your sample and set the parameters for the measurement.
2. Add a blank solution to a cuvette and ensure that the filling volume is sufficient to allow light path to pass through the solution.
3. Insert cuvette into the cell holder. 4. Initiate a blank measurement with the blank button. Upon completion of measurement
remove the cuvette. 5. Add sample to a cuvette and ensure that the sample volume is sufficient to allow light to
pass through the sample. 6. It is possible to enter a sample name for each sample in the input window “enter sample
name”. Note: Allowed characters are: A…Z a…z 0…9 , . – ( ) @ ! = _ ~ ; [ ] { } ` blank character 7. Initiate a sample measurement with the sample button. Upon completion of measurement remove the cuvette. 8. Apply further samples.
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SAMPLE HANDLING TIPS
NanoVolume Methods (N50-Go)
The sample window on pedestal is illuminated (N50-Go) with a low energy red light to assist with accurate sample application. The red light is switched off once the lid arm is closed. It is possible to disable the illumination feature in preferences of the NPOS software.
The minimum volume that can be used for NanoVolume samples is 0.3 µl (N50-Go dsDNA > 420 ng/µl and BSA > 12.6 mg/ml). For automatic path length setting at least 1µl is needed.
The maximum volume that can be used for NanoVolume samples is 2.0 µl (N50-Go). The sample can be fully recovered after measurement with a pipette if desired.
Note: Minimal cross contamination cannot be avoided on molecular level. Proper cleaning is important to ensure accurate measurements. In most cases a dry lint-
free laboratory wipe is sufficient to clean the sample quartz surfaces. In the case of highly concentrated samples or certain proteins, the recommended procedure for cleaning is to use a slightly wet lint-free laboratory wipe (with water or 70% EtOH depending on sample type) to thoroughly clean the sample surface. It is mandatory that the metal contact face around the measurement window and the mirror is clean. Cuvette Methods (C40-Go)
The cuvette holder is compatible with standard 10 mm path length quartz, glass and plastic cuvettes with an optical height of 8.5 mm.
It is also possible to use cuvettes with 5 mm, 2 mm, 1 mm or 0.5 mm path lengths, but there may be an adapter necessary. Please ask your cuvette supplier for a suitable adapter.
The minimum volume for accurate measurements depends on the cuvette type used; it is necessary that the light passes through the sample for accurate measurements. Center height is 8.5 mm.
Note: The cuvette holder is not removable. Do not pour any cleaning solution into the cuvette holder as larger amounts of liquids can get into the instrument and cause damage.
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SOLVENT COMPATIBILITY (N50-GO)
Most solvents typically used in life science laboratories are compatible with the NanoPhotometer® Go NanoVolume sample surfaces. The following solvents are compatible for use with the NanoPhotometer® models N50-Go at room temperature:

Acetone ( 5%)

Methylene chloride

Acetonitrile

MOPS

Benzene

Phenol (1%)

Butanol

N-propanol

Carbon tetrachloride

Toluene

Chloroform

Phosphate containing buffers

Ethanol

PBS (pH 4-10)

Ether

Citrate

HEPES

Borate

Hexane Isopropanol MES

Chloride salts
Acids > pH 2 Bases < pH 10

Methanol

Note: Highly concentrated acids and bases are not recommended. It is recommended to

wipe the sample surface with a lint-free laboratory wipe immediately upon completion of

each measurement. For more information about compatibility of specific solvents not listed

above, please contact the Implen support team (support@implen-go.com) to check the

compatibility.

DATA TRANSFER VIA FILE SERVER
All data saved on the NanoPhotometer® Go can easily be accessed from and transferred to a computer via the NanoPhotometer® Go file server. Connection options are LAN/WLAN, USB cable or WiFi Hotspot. It is possible to create user accounts for password secured file server access. User accounts for file server access can be activated in Preferences see page 94 File Server Access.
File Server Access via LAN/WLAN
For the file server access via LAN/WLAN it is necessary that both the computer and the NanoPhotometer® Go are connected to the same LAN/WLAN network. For connection of the NanoPhotometer® Go to LAN/WLAN see page 92 Network.

For Windows computer open the Windows explorer and enter the serial number or the NanoPhotometer® Go IP in the address bar of the Windows Explorer (e.g. \M80798 or
\Assigned IP Address).

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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2 Note: Serial number and IP address of the NanoPhotometer® Go can be found in the NanoPhotometer® Go software under Preferences/General/About. For a MAC computer open the “Connect to Server” dialog in the “Go” menu of the Mac OS X Finder and enter the NanoPhotometer® Go serial number or the active NanoPhotometer® Go IP address in the server address field to connect.
Note: Serial number and IP address of the NanoPhotometer® Go can be found in the NanoPhotometer® Go software under Preferences/General/About.
File Server Access via USB cable For file server access via USB cable connection, connect the NanoPhotometer® Go with a USB A/B cable to the computer and open the Windows Explorer or Connect to Server option for Mac (see file server access via LAN/WLAN) and enter \192.168.7.1 for connection.
File Server Access via WiFi Hotspot For file server access via WiFi Hotpot the WiFi Hotspot needs to be active on the NanoPhotometer® Go. For activation see page 93
WLAN Settings. The computer needs to be connected to the NanoPhotometer® Go WiFi Hotspot (SSID: NanoPhotometer® Go serial number; password: Implenuser). Open the Windows Explorer or Connect to Server option for Mac (see file server access via LAN/WLAN) and enter \192.168.8.1 for connection.
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4. NANOPHOTOMETER® GO APPLICATIONS
The NanoPhotometer® Go comes with pre-programmed applications. For the N50-Go are only the Nucleic Acid und Protein UV method available. There is an N50 upgrade option available for full applicational access. Any application method can be selected by tapping the icon once or clicking on the icon (computer based software).
NUCLEIC ACIDS
METHOD OVERVIEW
Nucleic acids in solution absorb light with a peak in the ultraviolet region of 260 nm. For determination of nucleic acid concentration in solution the absorbance at wavelength 260 nm is used along with the Beer-Lambert law. In addition to calculating concentrations of nucleic acids, absorbance measurements are also useful for estimating purity of nucleic acids by calculating the 260/280 nm and 260/230 nm ratios. Further, it is possible to determine the degree of labeling of nucleic acids with probes including fluorescent dyes. Sample ControlTM gives useful information about sample conditions. It recognizes air bubbles, sample impurities, turbidity, lint residues and potential contaminations. If Sample ControlTM detects any inconstancy an alert icon is shown in the result/table area. A push on the alert icon shows additional information about the inconstancy.
MEASUREMENT PROTOCOL
1. Select the Nucleic Acids icon on the home screen.
2. To change the nucleic acid type push on dsDNA and a list with available options opens on the right side. Options are: dsDNA, ssDNA, RNA miRNA, miRNA Sequence, Oligo, Oligo Sequence and Custom (see Table 1 on page 43). It is possible to enter a name for the custom nucleic acid factor for documentation.
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2 3. For NanoVolume application (N50-Go):
Select the volume of sample to be applied.
Note: 1 – 2 µl (default): automatic path length setting; 0.3 µl measures only the 0.07 mm path length (for samples with concentrations > 420 ng/µl dsDNA) For cuvette application (C40-Go): Select the path length depending on the used cuvette. Options are: 0.5 mm, 1 mm, 2 mm, 5 mm and 10 mm
If it is desired to heat the sample to 37°C use the toggle switch to turn on cell holder heater. When the cuvette holder has reached 37°C the toggle color changes to green. Note: Only available for cuvette applications (C40-Go). 4. Select the Units in which the concentration should be calculated. Options are ng/µl (default), µg/µl and µg/ml. pmol/µl if a nucleic acid sequence is entered for nucleic acid factor calculation.
5. The background correction is enabled at 320 nm by default. Selection options are 320 nm, 340 nm or any wavelength in the range of 220 ­ 350 nm. The background correction can be disabled with the toggle switch.
6. Air bubble recognition is disabled by default. Setting to on detects air bubbles, lint residues and bad sample conditions of the sample. Note: Lint residues and bad sample conditions are detected even if the air bubble recognition is set to off.
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2 7. For dye labelled samples tick the dye labels in the list, which should be used for result
calculation. Note: If the used dye is not available in the list, please go to preferences and add a custom dye to the dye list. There is an option for dye correction which can be enabled/disabled with a toggle switch. Note: Dye correction is only available for single dye selection.
8. Option to set/calculate a dilution factor for manual diluted samples.
9. Apply the blank ddH20 or buffer to the illuminated sample window on the pedestal and close the lid arm for the reference measurement and select blank to initiate the reading. Note: The illumination of the sample window can be switched off in the preferences.
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10. Use a lint-free laboratory wipe to clean both the sample window on pedestal and mirror in lid arm prior to applying the next sample.
Note: It could be helpful to apply the blank a second time and read it as a sample to ensure a proper blank.

11. Apply sample to the sample window on pedestal and press the sample button to initiate the measurement.

CALCULATIONS
Nucleic Acid Concentration
To determine the concentration of nucleic acids in solution, the absorbance is measured at a wavelength of 260 nm. The function describing the relationship between concentration and absorbance is a modification of the Beer-Lambert law equation. The concentration of the nucleic acid samples can be calculated with or without background correction depending on enabled/disabled background correction parameter.

Without background correction:

C = A260 Factornuc With background correction: C = (A260 – ABKG) Factornuc

C

Concentration in ng/µl

A260 ABKG

Absorbance at 260 nm (10 mm path) Absorbance at selected background wavelength (10 mm path) Manual dilution factor

Factornuc Nucleic acid factor in ng*cm/µl

Table 1. Nucleic acids extinction coefficients (nuc)

Type dsDNA ssDNA RNA miRNA Oligo miRNA Seq. Oligo Seq. Custom

Factornuc 50 ngcm/µl 37 ngcm/µl 40 ngcm/µl 33 ngcm/µl 33 ngcm/µl calculated via extinction coefficient of constituent nucleotides entered calculated via extinction coefficient of constituent nucleotides entered Option to enter any factor between 15 and 150 ngcm/µl

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Dye-labeled Nucleic Acid Concentration For dye-labeled nucleic acids, the concentration of the nucleic acid is calculated using a modified form of the Beer-Lambert equation. For these calculations, the instrument considers the absorption maximum of the dye and a certain dye-specific correction factor at 260 nm (see Table 2 on page 46). The concentration of a dye-labeled nucleic acid is calculated with or without background/dye correction as follows:
With background and with dye correction:
C = [(A260 – ABKG) – (cfdye (Amax, dye – ABKG))] Factornuc
With background and without dye correction: C = (A260 – ABKG)
Factornuc Without background and with dye correction: C = [A260 – (cfdye Amax, dye)] Factornuc Without background and without dye correction:
C = A260 Factornuc

C

Concentration in ng/µl

A260

Absorbance at 260 nm (10 mm path)

ABKG Absorbance at selected background wavelength (10 mm path)

Amax, dye Absorbance value at the absorbance maximum of the dye (10 mm path)

Factornuc Nucleic acid factor in ng*cm/µl

cfdye

Dye-dependent correction factor at 260 nm

Manual dilution factor

Note: Dye correction is only available for single dye selection.

Dye Concentration
For dye-labeled nucleic acids, the concentration of the dye is calculated using a modified form of the Beer-Lambert equation. For these calculations, the instrument considers the absorption maximum of the dye, the dye-specific extinction coefficient (see Table 2 on page 46). The dye concentration is calculated with or without background correction as follows:

With background correction:

C

=

(Amax, dye – dye*

ABKG) 10-6

Ð

Without background correction:

C

=

Amax, dye mdye 10-6

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C Amax, dye ABKG mdye

Concentration in ng/µl Absorbance value at the absorbance maximum of the dye (10 mm path)
Absorbance at 320 nm (10 mm path) Molar extinction coefficient of dye in M-1*cm-1
Manual dilution factor

Frequency of Incorporation (FOI)
FOI is the degree of labeling based on dye incorporation in a labeled nucleic acid sample. It is generally expressed as the number of dye molecules incorporated per 1000 nucleotides. FOI can be calculated with or without background/dye correction as follows:

With background correction with dye correction:

FOI =

324.5 * (Amax, dye – ABKG)

dye 10-6 (A260 – ABKG – cfdye (Amax, dye – ABKG)) Factornuc

With background correction without dye correction:

FOI

=

324.5 mdye 10-6

  • (Amax, dye – ABKG) (A260 – ABKG) * Factornuc

Without background correction and with dye correction:

FOI

=

mdye *

324.5 Amax, dye 10-6 (A260 – cfdye * Amax, dye)

  • Factornuc

Without background correction and without dye correction:

FOI

=

324.5 Amax, dye mdye 10-6 A260 Factornuc

FOI
A260 ABKG Amax, dye mdye
Factornuc cfdye

Frequency of Incorporation (dye per 1,000 bases) Absorbance at 260 nm (10 mm path) Absorbance at selected background wavelength (10 mm path) Absorbance value at the absorbance maximum of the dye (10 mm path) Extinction coefficient of dye in M-1cm-1
Nucleic acid factor in ng
cm/µl Dye-dependent correction factor at 260 nm

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Table 2. Dye Types, Absorbance Max, Extinction coefficient, and dye-dependent correction factors

NanoPhotometer® Go Model
N50-Go/C40-Go N50-Go/C40-Go N50-Go/C40-Go N50-Go/C40-Go N50-Go/C40-Go N50-Go/C40-Go N50-Go/C40-Go C40-Go C40-Go C40-Go N50-Go/C40-Go N50-Go/C40-Go C40-Go C40-Go N50-Go/C40-Go N50-Go/C40-Go N50-Go/C40-Go C40-Go C40-Go C40-Go N50-Go/C40-Go

Dye Type

Absorbance maximum of Dye (nm)

Molar ext. coeff.
of dye mdye in M-1*cm-1

Alexa Fluor 350

346

Alexa Fluor 488

495

Alexa Fluor 532

532

Alexa Fluor 546

554

Alexa Fluor 555

555

Alexa Fluor 568

578

Alexa Fluor 594

590

Alexa Fluor 647

650

Alexa Fluor 660

663

Alexa Fluor 680

679

Cy3

550

Cy3.5

581

Cy5

649

Cy5.5

675

Oyster-500

503

Oyster-550

553

Oyster-556

560

Oyster-645

649

Oyster-650

653

Oyster-656

660

Texas Red

603

19,000 71,000 81,000 112,000 150,000 91,300 90,000 239,000 132,000 184,000 150,000 150,000 250,000 250,000 78,000 150,000 155,000 220,000 200,000 200,000 112,000

Dye-dependent correction factor at 260 nm cfdye
0.25 0.30 0.24 0.21 0.08 0.45 0.43 0.00 0.00 0.00 0.08 0.08 0.05 0.05 0.29 0.05 0.03 0.05 0.04 0.04 0.23

Ratios
Reactions utilizing nucleic acids often require minimum purity standards. Common contaminants of nucleic acid samples include: proteins, organic compounds, and other. Based on the common contaminants of nucleic acid samples, the 260/280 and 260/230 ratios are calculated for nucleic acids to give an indication of the purity of the samples. Pure DNA and RNA preparations have expected 260/280 ratios of 1.8 and 2.0 respectively. An elevated absorbance at 230 nm can indicate the presence of impurities as well; 230 nm is near the absorbance maximum of peptide bonds and also indicates buffer contamination since TRIS, EDTA and other buffer salts absorb at 230nm. When measuring RNA samples, the 260/230 ratio should be > 2.0; a ratio lower than this is generally indicative of contamination with guanidinium thiocyanate, a reagent commonly used in RNA purification and which absorbs over the 230-260 nm range. If a ratio is detected out of the acceptable
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2 range an alert icon is shown in the results/table area. A push on the alert icon shows additional information. The ranges for acceptable ratio values can be defined in preferences. The ratios are calculated with or without background correction according to if the background correction is activated during the measurements or not as follows: Without background correction: 260/280 ratio = A260
A280
260/230 ratio = A260
A230
With background correction: 260/280 ratio = A260 – ABKG
A280 – ABKG
260/230 ratio = A260 – ABKG
A230 – ABKG
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PROTEIN UV
METHOD OVERVIEW
The Protein UV method exploits the inherent absorbance of proteins at 280 nm in combination with the Beer-Lambert Law, where each protein is characterized by a protein specific extinction coefficient () which can be used to determine total protein concentration of a solution. The intrinsic absorbance of proteins is due to the presence of aromatic amino acids in their structure, primarily tryptophan and tyrosine, as well as cysteine (oxidized cysteine residues in a disulphide bond). The aromatic amino acid residues in a protein containing tryptophan and tyrosine exhibit strong intrinsic absorbance at 280 nm, with a lesser contribution by phenylalanine. Therefore, it is the aromatic amino acid residues which dictate the extinction coefficient at 280 nm for a protein.
The most straightforward method to determine concentration of a purified, homogenous protein with a known extinction coefficient () is by direct measurement of UV280 provided as long as the protein contains no prosthetic groups with strong absorption in the same region. However, for unknown proteins including homogenous protein mixtures, it is possible to make direct A280 measurements using a composite value derived from comparison of many proteins, although this will only provide an approximate but close estimate of the true protein concentration.
The NanoPhotometer® Go determines protein concentration by performing calculations based on specific values, either pre-programmed in the instrument or entered manually by the user. Extinction coefficient () values at 280 nm vary greatly for different proteins due to their particular aromatic amino acid content. Fixed values are pre-programmed in the software for certain proteins (see Table 3 on page 52). However, if the protein of interest is not included in the pre-programed methods it is also possible to manually enter the specific for the protein of interest using the custom Mol. Ext. Coefficient, custom Ext. Coefficient or custom 1/ protein factor option. For correct calculation, it is necessary to supply either: a) the molar extinction coefficient (M in M-1cm-1) and the molecular weight expressed in molar mass units (g/mol); b) the mass extinction coefficient ( in l/gcm) or c) the protein factor 1/ of the protein.
To determine the degree of dye labelling of a protein, the absorbance measured at the wavelength corresponding to the absorbance maximum of the fluorescence dye is used (see Table 3 on page 52). The corresponding extinction coefficient of the dye is used along with the Beer-Lambert Law to determine the dye concentration.
Note: It is important to ensure the extinction coefficient and units entered are correct in order to ensure that calculations are performed properly for accurate concentration values.
Sample ControlTM gives useful information about air bubbles, sample impurities, turbidity, lint residues and potential contaminations. If the Sample ControlTM detects any inconstancy an alert icon is shown in the result/table area. A push on the alert icon shows additional information about the inconstancy.
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MEASUREMENT PROTOCOL
1. Select the Protein UV icon on the home screen.
2. To change the protein type push on BSA and a list with available options opens on the right side. Options are: BSA, SA Mouse, SA Human, IgG Mouse, IgG Human, IgE Human, Lysozyme, OD1, Custom (Molar Extinction Coefficient), Custom (Extinction Coefficient) and Custom (1/). For Custom (Mol. Ext. Coefficient) enter Molecular Weight in g/mol and the Mol. Ext. Coefficient in M-1cm-1 For Custom (Ext. Coefficient) enter the Ext. Coefficient in l/gcm For Custom (1/) enter the calculated protein factor 1/
3. For NanoVolume application (N50-Go):
Select the volume of sample to be applied.
Note: 1-2 µl (default): automatic path length change; 0.3 µl measures only the 0.07 mm path length (possible for samples with concentrations e.g. BSA > 12.6 mg/ml) For cuvette application (C40-Go): Select the path length depending on the used cuvette. Options are: 0.5 mm, 1 mm, 2 mm, 5 mm and 10 mm
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If it is desired to heat the sample to 37°C use the toggle switch to turn on cell holder heater. When the cuvette holder has reached 37°C the toggle color changes to green. Note: Only available for cuvette applications (C40-Go) 4. The wavelength for protein measurements can be changed in the range of 200 ­ 330nm depending on the wavelength peak of the protein. Default setting is 280 nm 5. Select the Units in which the concentration should be calculated. Options are ng/µl, µg/µl, µg/ml and mg/ml (default).
6. The background correction is enabled at 320 nm by default. Selection options are 320 nm, 340 nm or any wavelength in the range of 220 ­ 350 nm. The background correction can be disabled with the toggle switch.
7. Air bubble recognition is disabled by default. When enabled it detects air bubbles, lint residues and poor conditions of the sample. Note: Lint residues and bad sample conditions are detected even if the air bubble recognition is set to off.
8. For dye labelled samples, tick the dye labels in the list which should be used for result calculation. Note: If the used dye is not available in the list, please go to preferences and add a custom dye to the dye list. 50

NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2 There is an option for dye correction which can be enabled/disabled with a toggle switch.
Note: Dye correction is only available for single dye selection.
9. Option to set/calculate a dilution factor for manual diluted samples.
10. Apply the blank ddH20 or buffer to the illuminated sample window on pedestal for the reference measurement and select blank to initiate the reading. Note: The illumination of the sample window can be switched off in the preferences.
11. Use a lint-free laboratory wipe to clean both the sample window on pedestal and mirror in the lid arm prior to applying the next sample. Note: It can be helpful to apply the blank a second time and read it as a sample to ensure a proper blank.
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12. Apply sample to the sample window on pedestal and press the sample button to initiate the measurement.

CALCULATIONS
Protein UV280 Concentration
The protein concentration in the Protein UV method is calculated with the absorbance value of the sample at 280 nm or entered wavelength in the range of 220-350 nm along with the extinction coefficient defined by the user. The protein concentration is calculated with or without background correction as follows:

With background correction: C = (A280 – ABKG) Factorprot Ð

Without background correction:

C = A280 Factorprot Ð

C

Concentration (mg/ml)

A280

Absorbance at 280 nm (10 mm path) or entered wavelength

ABKG Absorbance at selected background wavelength (10 mm path)

Factorprot Protein factor in g*cm/l (1/Ext. Coeff. or MW/Mol.Ext. Coeff.)

Ð

Dilution factor

Table 3. Protein extinction coefficients (prot)

Type BSA SA Mouse SA Human IgG Mouse IgG Human IgE Human Lysozyme OD1

Factorprot [ g* cm/l] 1.499 1.493 1.718 0.714 0.735 0.654 0.379 1.000

Ext. Coeff. [l/g*cm] 0.6670 0.6700 0.5820 1.4000 1.3600 1.5300 2.6400 N/A

Mol. Ext. Coeff. [M-1*cm-1 ] 44,289 44,220 40,370 224,000 204,000 290,700 37,984 N/A

MW [g/mol] 66,400 66,000 69,365 160,000 150,000 190,000 14,388 N/A

Dye-labeled Protein UV280 Concentration
For dye-labeled proteins, the concentration of the protein is calculated using a modified form of the Beer-Lambert equation. For these calculations, the instrument considers the absorption maximum of the dye, and a certain dye- specific correction factor at 280 nm (see Table 4 on page 54). The dye concentration is calculated with or without background/dye correction for are as follows:
With background and with dye correction:
C = [A280 – ABKG – (cfdye (Amax, dye – ABKG))] Factorprot * Ð

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With background and without dye correction: C = (A280 – ABKG) Factorprot Ð Without background and with dye correction:

C = (A280 – ( cfdye Amax, dye)) Factorprot * Ð

Without background and without dye correction:

C = A280 Factorprot Ð

C
A280 ABKG Amax, dye Factorprot cfdye Ð

Concentration in mg/ml Absorbance at 280 nm (10 mm path) Absorbance at selected background wavelength (10 mm path) Absorbance value at the absorbance maximum of the dye (10 mm path) Protein factor in g*cm/l (1/Ext. Coeff. or MW/Mol.Ext. Coeff.) Dye-dependent correction factor at 280 nm Dilution factor

Dye Concentration

For dye-labeled proteins, the concentration of the dye is calculated using a modified form of the Beer-Lambert equation. For these calculations, the instrument considers the absorption maximum of the dye, and a dye-specific extinction coefficient (see Table 4 on page 54). The dye concentration is calculated with or without background correction for are as follows:

With background correction:

C

=

((Amax, dye – ABKG) mdye * 10-6

  • Ð)

Without background correction:

C

=

(Amax, dye Ð) dye 10-6

C Amax, dye ABKG mdye Ð

Concentration Absorbance at the max dye absorbance value (10 mm path)
Absorbance at selected background wavelength (10 mm path) Extinction coefficient of dye in M-1*cm-1
Dilution factor

Degree of Labeling (DOL)
DOL is the degree of labeling based on the average number of dye molecules coupled to a protein molecule. The degree of labeling can be determined from the absorption spectrum of the labeled antibody with or without background/dye correction as follows:

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With background and with dye correction:

DOL =

(Amax, dye – ABKG) * mprot

((A280 – ABKG) – cfdye * (Amax, dye – ABKG)) mdye

With background and w/o dye correction:

DOL =

(Amax, dye (A280 –

– ABKG mprot ABKG) mdye

W/o background and with dye correction:

DOL =

(A280

Amax, dye mprot – cfdye Amax, dye) * mdye

W/o background and w/o dye correction:

DOL =

Amax, dye mprot A280 mdye

DOL
Amax, dye
A280 ABKG mdye mprot cfdye

Degree of labeling/dye per protein ratio Absorbance value at the absorbance maximum of the dye (10 mm path)
Absorbance at 280 nm (10 mm path) Absorbance at selected background wavelength (10 mm path) Extinction coefficient of dye in M-1cm-1 Molar extinction coefficient of protein (M-1 cm-1)
Dye-dependent correction factor at 280 nm

Table 4. Dye Types, Absorbance Max, Extinction coefficients, and dye-dependent correction factors

NanoPhotometer® Go Dye Type Models

N50-Go/C40-Go N50-Go/C40-Go N50-Go/C40-Go N50-Go/C40-Go N50-Go/C40-Go N50-Go/C40-Go N50-Go/C40-Go N50-Go/C40-Go C40-Go C40-Go C40-Go N50-Go/C40-Go C40-Go C40-Go

Alexa Fluor 350 Alexa Fluor 405 Alexa Fluor 488 Alexa Fluor 532 Alexa Fluor 546 Alexa Fluor 555 Alexa Fluor 568 Alexa Fluor 594 Alexa Fluor 647 Alexa Fluor 680 Alexa Fluor 790 Cy3 Cy5 DyLight 649

Absorbance maximum of Dye (nm)
346 401 495 532 554 555 578 590 650 679 785 550 649 654

Molar ext. coeff. of Dye mdye in M-1*cm-1
19,000 34,000 71,000 81,000 112,000 150,000 91,300 90,000 239,000 184,000 260,000 150,000 250,000 250,000

Dye-dependent correction factor at 280 nm cfdye
0.19 0.70 0.11 0.09 0.12 0.08 0.46 0.56 0.03 0.05 0.08 0.05 0.05 0.04

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N50-Go/C40-Go

DyLight 488

493

70,000

0.15

N50-Go/C40-Go

FITC

494

70,000

0.30

N50-Go/C40-Go

Pacific Blue

409

30,000

0.20

N50-Go/C40-Go

Pacific Orange

397

24,500

0.60

N50-Go/C40-Go

pHrodo Green

505

75,000

0.20

N50-Go/C40-Go

pHrodo Red

560

65,000

0.12

N50-Go/C40-Go

r-PE

566

1,863,000

0.17

N50-Go/C40-Go

Texas Red

595

80,000

0.18

Ratios
Protein samples e.g. from whole cell lysates may contain nucleic acids. To check the purity of the isolated protein, the 260/280 ratio is calculated to give an indication of the nucleic acid contamination. A pure protein preparation has an expected 260/280 ratios of 0.57. If a ratio is detected out of the acceptable range an alert icon is shown in the results/table area. A push on the alert icon shows additional information. The ranges for acceptable ratio values can be defined in preferences. The ratio is calculated with or without background correction according to if the background correction is activated during the measurements or not as follows:
Without background correction:
260/280 ratio = A260
A280
With background correction:
260/280 ratio = A260 – ABKG
A280 – ABKG

PROTEIN ASSAYS
Available only for C40-Go, N50-Go needs upgrade for activation of Protein Assays.
METHOD OVERVIEW
Protein concentration may be measured using colorimetric assays, in which certain reagents are added to the protein solution to generate a colored product; either a protein-cupric ion chelate as in the Biuret, Lowry, BCA assays or a protein-dye complex as in the Bradford assay. In these colorimetric assays, the absorbance is measured in the visible range at the appropriate wavelength for each assay and compared against a standard curve prepared by serial dilution of a protein standard of known concentration. A linear, linear zero or 2nd order regression analysis of the calibration standard data points is calculated by the NanoPhotometer® Go. A correlation coefficient (R2) in the range of 0.95 to 1.00 indicates a good fit to a straight line.
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Bradford Assay Method depends on quantifying the binding of a dye, Coomassie Brilliant Blue, to an unknown protein and comparing this binding to that of a standard curve prepared from a set of known protein of known concentrations at 595 nm. This standard is usually BSA (bovine serum albumin).
Biuret Assay Method depends on a reaction between cupric ions and peptide bonds in an alkali solution, resulting in the formation of a complex absorbing at 546 nm.
BCA Assay Method depends on a reaction between cupric ions and peptide bonds coupled with the detection of cuprous ions using bicinchoninic acid (BCA), giving an absorbance maximum at 562 nm. The BCA process is less sensitive to the presence of detergents used to solubilize membranes.
Lowry Assay Method is based on the Biuret reaction. Under alkaline conditions the divalent copper ion forms a complex with peptide bonds in which it is reduced to a monovalent ion. Monovalent copper ion and the radical groups of tyrosine, tryptophan, and cysteine react with Folin reagent to produce an unstable product that becomes reduced to molybdenum/tungsten blue. The bound reagent changes color from yellow to blue. This binding is compared with that obtained with a standard protein at 750 nm; this is usually BSA (bovine serum albumin). Note: Detailed protocols are customarily supplied with these assay kits, and must be closely followed to ensure that accurate results are obtained.
MEASUREMENT PROTOCOL
1. Select protein assays icon from home screen.
2. To change the assay type click on Bradford and a list with available options opens on the right side. Options are: BCA Assay, Biuret Assay (not for N50 Go), Bradford Assay, Lowry Assay (not for N50-Go)
3. For Cuvette Application (C40-Go): Select the path length depending on the used cuvette. Options are: 0.5 mm, 1 mm, 2 mm, 5 mm and 10 mm
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If it is desired to heat the sample to 37°C use the toggle switch to turn on cell holder heater. When the cuvette holder has reached 37°C the toggle color changes to green. Note: Only available for cuvette applicatons (C40-Go). For NanoVolume Application (only with N50-Go Upgrade): Select the dilution depending on the sample concentration
Note: There is no automatic path length setting available in this method. Select either a virtual dilution of 15 (path length 0.67 mm) or of 140 (path length 0.07 mm) depending on your sample concentration. 4. Default values for the baseline correction are depending on selected protein assay type and instrument version: BCA default value 750 nm (N50: off) Biuret default off Bradford default value 750 nm (N50: 350 nm) Lowry default value 405 nm
Note: It is recommended to use the default baseline for each assay. 5. Select curve fit type: Options are linear regression, zero regression (forces the straight line
through the origin) and 2nd order regression.
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7. Add up to 20 Concentrations by pushing on the Add Concentration button. Added concentrations can be deleted with Enter the concentrations of the standards for the standard curve.
8. Select Replicates none, 2 or 3
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9. Measure a blank and depending on the replicate selection, all required concentrations. Absorbances of the replicates will be shown in the results area and if replicates are selected a mean value for each standard. It is possible to exclude single measurements from the curve calculation by switching the toggle switch off. Note: After the first sample measurement the standard curve can no longer be altered. Once a Standard curve is created or loaded from a stored method it will be used for concentration calculations of measured samples. It may be necessary to do a blank measurement.
10. Apply sample and press the sample button to initiate the sample measurement. Note: Once the sample measurement is initiated it is no longer possible to change the standard curve.
SAVING AND LOADING STANDARD CURVES
It is possible to save measured standard curves as a Stored Method. To save the standard curve push the store method button and enter a method name, select a folder and save with the Store button. Methods can be opened in the Stored Methods menu on the homescreen. Opening a saved Protein Assay method shows a message with the option to load or remeasure the standard curve.
CALCULATIONS
Protein concentration is determined using the standard curve by correlating absorbance values of samples with known concentration to calculate the concentration of the unknown sample. In order to maintain accuracy and precision please ensure that the R2 value of the standard curve is 0.95 or greater.
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KINETICS
Available only for C40-Go, N50-Go needs upgrade for activation of Kinetics.
METHOD OVERVIEW
The Kinetics application is useful for: measuring initial rates of enzyme- catalyzed reactions, performing progress-curve analysis for complete reactions, calculating basic MichaelisMenten parameters for single-substrate reactions, and measuring enzyme inhibition. Simple kinetics studies, where the change in absorbance is followed as a function of time at a fixed wavelength, can be readily performed with the NanoPhotometer® Go. The rate of a chemical reaction can be measured using spectrophotometric methods by studying the change in absorbance at a fixed wavelength as a function of time. These changes in absorbance reflect corresponding changes in the concentration of reactants or products as the reaction progresses. The rate of many chemical reactions can be markedly accelerated by the presence of catalysts, which remain chemically intact during the reaction. Catalysts in the case of biochemical reactions are generally represented by enzymes, which are specialized protein catalysts. However, a few examples of special reactions catalyzed by RNA molecules also exist. Studying the kinetics of a reaction can reveal important details of the catalytic mechanism involved in terms of sequence steps, transition state of reactants or nature of enzyme inhibitors.

MEASUREMENT PROTOCOL

Note: If a kinetic is started from a control device via WiFi connection (tablet or smartphone) set the auto lock of the tablet or smartphone to never. Otherwise the kinetics will be interrupted when the smartphone or tablet is locked, because of losing the WiFi connection.
1. Select the Kinetics icon on the home screen
2. For Cuvette Application (C40-Go): Select the path length depending on the used cuvette. Options are: 0.5 mm, 1 mm, 2 mm, 5 mm and 10 mm
If it is desired to heat the sample to 37°C use the toggle switch to turn on the cell holder heater. When the cuvette holder has reached 37°C the toggle color changes to green. Note: Only available for cuvette applications (C40-Go).
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For NanoVolume Application (only with N50-Go Upgrade): Select the dilution depending on the sample concentration.
Note: There is no automatic path length setting available in this method. Select either a virtual dilution of 15 (path length 0.67 mm) or of 140 (path length 0.07 mm). 3. Default wavelength () is 340 nm but can be changed in the range of 200­900 nm (N50: 200­650 nm), depending on the application. 4. Time settings: Enter the duration time in minutes over which measurements are to be taken. Possible
range is 1­3000 minutes. Enter the interval time between measurements in seconds. Possible interval times are 5-
3,600 seconds (N50: 10­3,600 sec.), depending on the duration time. Enter the delay time in seconds before the first measurement is taken. Possible delay
time is between 0­3,600 seconds, depending on the duration time.
Note: A maximum of 500 samples is possible. Please consider this when choosing the duration and interval time. 5. Insert cuvette with the reference sample and select blank button to initiate the measurement. 6. Insert cuvette with the sample and select the sample button to initiate the measurements. Once the kinetic is started the Blank button turns to a Pause/Continue button and the Sample button to a Stop button. Note: While the kinetics is running it is not possible to change the parameters, save data or delete data. Change of parameters is only possible before starting the kinetic readings. Save and delete data is only available after the kinetic session is stopped. Note: Auto print and cryo label print is not available in Kinetics.
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CALCULATIONS
All absorbance values are normalized to a 10 mm path.

A0 An dA Slope Final A
R2

= absorbance of start value (10 mm path)

= absorbance of actual value time n (10 mm path)

= absorbance of actual value ­ absorbance of start value

= linear regression fit of all actual measurement points

= absorbance of final value

=

R2

=

ni=1

(Oi -Ei)2 Ei

[Oi

=

observed

slope

value;

Ei

=

expected

slope

value]

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OD600
Available only for C40-Go, N50-Go needs upgrade for activation of OD600.
METHOD OVERVIEW
The growth of bacteria in liquid culture media is commonly monitored by measuring the optical density at 600 nm (OD600) in small samples taken from the cultures. OD600 measurements are typically used to determine the stage of growth of the bacterial culture, thereby ensuring that cells are harvested at an optimum point that corresponds to an appropriate density of live cells. Growth of bacterial cells typically progresses through a series of consecutive phases including: lag, log, stationary and decline (see Figure 1 on page 64). In general, cells should be harvested towards the end of the log phase using the optical density of the samples to determine when this point has been reached. Since optical density in the case of OD600 measurements results from light scattering rather than light absorption, this value varies depending on the type of bacterial cells in the culture in terms of size and shape. Cells are routinely grown until the absorbance at 600 nm (known as OD 600) reaches approximately 0.4 prior to induction or harvesting. A linear relationship exists between cell number (density) and OD 600 up to an absorbance value of 0.6, approximately.
As mentioned above, for turbid samples such as cell cultures, the absorbance measured is due to light scattering, and not the result of molecular absorption. Since the extent of scattering is affected by the optics of the system (distance between the cell holder and instrument exit slit, monochromator optics, slit geometry, etc.), different spectrophotometer types will tend to give different OD 600 readings for the same turbid sample. Therefore, if results from different spectrophotometers are to be compared, they must be normalized first using appropriate calibration curves. For more information see Technical Note #8 OD 600 which can be downloaded on the Implen webpage: www.implen-go.com/scientificpublications/
A calibration curve can be constructed by comparing measured OD 600 to expected OD 600. Expected OD 600 is determined by counting cell number using an alternative technique (for example microscope slide method) and converting to OD 600 using the rule of thumb that 1 OD 600 = 5 x 108 cells/ml for E. coli.
The NanoPhotometer® Go comes with a correction factor of 1 by default. To compare OD 600 values between different spectrophotometers, it is necessary to determine the constant deviation or ratio between the absorbance values for the same sample from each instrument and use this factor within the setting “correction factor” of your NanoPhotometer® Software.
Note: The use of 10 mm path length disposable cuvettes is recommended for optical density measurements of cell culture solutions. The amount of cells is reflected in the reading and the likelihood of fluctuating amount of cells in a drop from sample to sample can be considered as extremely significant. It is therefore recommended to use cuvettes since the amount of error in a bigger volume is not as significant. The cuvette measurements provide a bigger average and therefore more reproducible readings. Also, to prevent the suspension settling too quickly and giving an OD reading that changes with time, glycerol should be added to the sample.
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Figure 1 Bacterial growth curve
MEASUREMENT PROTOCOL
1. Select the OD600 icon on the home screen
2. For Cuvette Application (C40-Go): Select the path length depending on the used cuvette. Options are: 0.5 mm, 1 mm, 2 mm, 5 mm and 10 mm
If it is desired to heat the sample to 37°C use the toggle switch to turn on cell holder heater. When the cuvette holder has reached 37°C the toggle color changes to green. Note: Only available for cuvette applications (C40-Go). For NanoVolume Application (only with N50-Go Upgrade): Select the dilution depending on the sample concentration
Note: There is no automatic path length setting in this method. Select either a virtual dilution of 15 (path length 0.67 mm) or of 140 (path length 0.07 mm)
3. Default wavelength is 600 nm but the wavelength can be changed in the range of 200­900 nm (N50-Go: 200­650 nm), depending on the application.
4. Toggle switch cells/ml is disabled by default. Enable cells/ml to get the cells/ml calculated. Enter the cell specific factor and multiplier (e.g. 1 OD600 = 5 x 108 cells/ml) 64

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5. Enter the correction factor to compensate for different optical configurations between the NanoPhotometer® Go and other instruments.
6. Option to smooth the graph with different boxcars. Options: Off, 1 = boxcar 11 (default), 2 = boxcar 21 and 3 = boxcar 61
7. Option to set/calculate a dilution factor for manual diluted samples.
8. Insert cuvette with the reference sample and select blank button to initiate the measurement 65

NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2
9. Insert cuvette with the sample and select the sample button to initiate the measurement.

CALCULATIONS

OD600 = A600 Ð cf

OD600 A600 Ð cf

Optical density at 600 nm Absorbance at 600 nm (10 mm path) Dilution factor Correction factor for spectrophotometer

Cells/ml = A600 Ð cf * multiplier

A600 Ð cf multiplier

Absorbance at 600 nm (10 mm path) Dilution factor Correction factor for spectrophotometer Multiplier of sample

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MORE APPS
Available only for C40-Go, N50-Go needs upgrade for activation of More Apps. The More Apps icon located on the home screen opens another menu screen with access to icons for additional applications available on the NanoPhotometer® Go. The applications featured in this menu include: wavelength, concentration, wavescan, absorbance/ratio, standard curve and custom applications.
MORE APPS: WAVELENGTH
Available only for C40-Go, N50-Go needs upgrade for activation of Wavelength.
METHOD OVERVIEW
In the wavelength application it is possible to measure simple absorbance (A) and % transmittance (%Trans. / only in cuvette mode) of a sample at specific wavelengths. It is possible to add up to 20 different wavelengths. The wavelength method includes a calculation tool to define and calculate customer defined formulas.
MEASUREMENT PROTOCOL
1. Select the More Apps icon from the home screen and the Wavelength icon from the More Apps screen.
2. For cuvette application (C40-Go): Select the path length depending on the used cuvette. Options are: 0.5 mm, 1 mm, 2 mm, 5 mm and 10 mm
If it is desired to heat the sample to 37°C use the toggle switch to turn on cell holder heater. When the cuvette holder has reached 37°C the toggle color changes to green. Note: Only available for cuvette applications (C40-Go). For NanoVolume application (only with N50-Go Upgrade): Select the dilution depending on the sample concentration.
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Note: There is no automatic path length setting in this method. Select either a virtual dilution of 15 (path length 0.67 mm) or of 140 (path length 0.07 mm). 3. Enter desired wavelength () to be measured. It is possible to measure up to 20 wavelengths simultaneously. More wavelength () options can be added by selecting the Add Wavelength button. Added wavelength can be deleted with
4. Baseline correction is set off by default. Enabling the baseline corrections shows a list with different wavelength options: 377 nm, 604 nm, 650 nm, 770 nm (N/A N50-Go) and 823 nm (N/A N50-Go). Option to enter any wavelength between 200 nm and 900 nm (N50-Go: 650 nm).
5. Option to smooth the graph with different boxcars. Options: Off, 1 = boxcar 11 (default), 2 = boxcar 21 and 3 = boxcar 61
6. Option to enter formulas to calculate
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It is possible to enter up to 5 different custom formulas for result calculation. Formula Acceptance Guidelines: 1. Numbers:
Up to 20 significant figures if no decimal separator is used Up to 4 significant figures if a decimal separator (period “.”) is used 2. Numeric Operations: + (add), – (subtract), (multiply), / (divide) and parentheses () 3. Absorbance: Axxx e.g. for absorbance at 260 nm: A260 Note: Do not use blank character. Example: Nucleic Acid (dsDNA) concentration calculation with background correction at 320 nm: (A260-A320)50 7. Option to set/calculate a dilution factor for manual diluted samples.
8. Apply the blank ddH20 or buffer to the illuminated sample window on pedestal for the reference measurement and select blank to initiate the reading.
Note: The illumination of the sample window can be switched off in preferences.
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9. Use a lint-free laboratory wipe to clean both the sample window on pedestal and mirror in lid arm prior to applying the next sample. Note: It can be helpful to apply the blank a second time and read it as a sample to ensure a proper blank.

Apply sample to the sample window on pedestal and press the sample button to initiate the measurement.

CALCULATIONS
Formula Calculation: Is depending on the entered formula in the parameter concentration.

Absorbance Calculation:
Absorbance is formally defined as the decimal logarithm (base 10) of the reciprocal of transmittance:

A = log (T1) = – log T T = 10(-A)

Note: Corresponding absorbance value e.g. Absorbance value ( = 230) etc. normalized to 10 mm path length

%Transmittance Calculations (cuvette mode only)

In the wavelength application it is possible to measure the absorbance (A) and % transmittance (%T) of a sample with respect to a reference at a specific wavelength. Transmittance is the ratio of light intensity remaining after it has passed through the sample (I) to the initial incident light intensity (I0):

=

0

%

=

0

×

100

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MORE APPS: WAVESCAN
Available only for C40-Go, N50-Go needs upgrade for activation of Wavescan.
METHOD OVERVIEW
Using the wavescan application it is possible to obtain the complete spectral scan for a defined wavelength range between 200-900 nm (C40-Go) or from 200-650 nm (N50Go/Upgrade necessary).
MEASUREMENT PROTOCOL
1. Select the More Apps icon from the Home screen and the Wavescan icon from the More Apps screen
2. For cuvette application (C40-Go): Select the path length depending on the used cuvette. Options are: 0.5 mm, 1 mm, 2 mm, 5 mm and 10 mm
If it is desired to heat the sample to 37°C use the toggle switch to turn on cell holder heater. When the cuvette holder has reached 37°C the toggle color changes to green. Note: Only available for cuvette applications (C40-Go). For NanoVolume application (only with N50-Go Upgrade): Select the dilution depending on the sample concentration Note: There is no automatic path length setting in this method. Select either a virtual dilution of 15 (path length 0.67 mm) or of 140 (path length 0.07 mm)
3. Set Start and End Wavelength to define the scan range.
Note: If samples with a different wavelength range are selected, the graphs are shown on full scan range of 200­900 nm (N50-Go: 200-650 nm). 4. Baseline correction is set off by default. Enabling the baseline corrections shows a list with
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2 different wavelength options: 377 nm, 604 nm, 650 nm, 770 nm (N/A N50) and 823 nm (N/A N50). Option to enter any wavelength between 200 nm and 900 nm (N50: 650 nm)
5. Option to smooth the graph with different boxcars. Options: Off, 1 = boxcar 11 (default), 2 = boxcar 21 and 3 = boxcar 61
6. Option to set/calculate a dilution factor for manual diluted samples.
7. Apply the blank ddH20 or buffer to the illuminated sample window on pedestal for the reference measurement and select blank to initiate the reading. Note: The illumination of the sample window can be switched off in the preferences.
8. Use a lint-free laboratory wipe to clean both the sample window on pedestal and mirror in lid arm prior to applying the next sample. Note: It can be helpful to apply the blank a second time and read it as a sample to ensure a proper blank.
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button to initiate the measurement.
CALCULATIONS
No calculations necessary: values are reported based on 10 mm path length. The results show prominent peaks with wavelength and absorbance values. For cuvette measurements it is possible to change to the %Transmittance mode. If a peak of interest is not shown in the results the peak can be selected by pushing on the graph. The peak can then be added to the results by tapping on the Add Peak button in the pop-up window.
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MORE APPS: ABSORBANCE RATIO
Available only for C40-Go, N50-Go needs upgrade for activation of Absorbance Ratio.
METHOD OVERVIEW
In this mode, it is possible to determine simple absorbance ratios for a given sample by measuring the absorbance at two wavelengths specified in the parameters of the method relative to a blank.
MEASUREMENT PROTOCOL
1. Select the More Apps icon from the home screen and the Absorbance/Ratio icon from the More Apps screen
2. For cuvette application (C40-Go): Select the path length depending on the used cuvette. Options are: 0.5 mm, 1 mm, 2 mm, 5 mm and 10 mm
If it is desired to heat the sample to 37°C use the toggle switch to turn on cell holder heater. When the cuvette holder has reached 37°C the toggle color changes to green. Note: Only available for cuvette applications (C40-Go). For NanoVolume application (only with N50-Touch Upgrade): Select the dilution depending on the sample concentration
Note: There is no automatic path length setting in this method. Select either a virtual dilution of 15 (path length 0.67 mm) or of 140 (path length 0.07 mm). 3. Enter desired wavelengths ( 1-1 and 1-2) for ratio calculation. It is possible to measure up to 20 absorbance/ratios simultaneously. More wavelengths for ratio calculation can be added by selecting the Add Ratio button. Added ratios can be deleted with .
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NanoPhotometer® N50-Go/C40-Go User Manual Version 4.6.2 4. Baseline correction is set to off as default. Enabling the baseline corrections shows a list
with different wavelength options: 377 nm, 604 nm, 650 nm, 770 nm and 823 nm. Option to enter any wavelength between 200 nm and 900 nm (N50: 650 nm)
5. Option to smooth the graph with different boxcars. Options: Off, 1 = boxcar 11 (default), 2 = boxcar 21 and 3 = boxcar 61
6. Option to set/calculate a dilution factor for manual diluted samples.
7. Apply the blank ddH20 or buffer to the illuminated sample window on pedestal for the reference measurement and select blank to initiate the reading. Note: The illumination of the sample window can be switched off in the preferences.
8. Use a lint-free laboratory wipe to clean both the sample window on pedestal and mirror in lid arm prior to applying the next sample. Note: It can be helpful to apply the blank a second time and read it as a sample to ensure a proper blank.
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9. Apply sample to the sample window on pedestal and press the sample button to initiate the measurement.

CALCULATIONS

The absorbance ratio is calculated from the two path lengths specified by the user in the parameters.

1: 2=

1 2

1: 2 = Absorbance Ratio 1 = Absorbance 1 corresponding absorbance value 1 selected normalized to 10 mm path
2 = Absorbance 2 corresponding absorbance value 2 selected normalized to 10 mm path

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MORE APPS: CONCENTRATION
Available only for C40-Go, N50-Go needs upgrade for activation of Concentration.
METHOD OVERVIEW
In this mode, concentration can be calculated for a sample by determining the absorbance at a specific wavelength relative to a reference. The concentration is then obtained by multiplying the measured absorbance by a specific factor. This factor may be known in advance and entered by the user, or it may be calculated by the instrument by measuring a set of standard (standard curve method) with known concentrations to create a standard curve.
MEASUREMENT PROTOCOL
1. Select the More Apps icon from the home screen and the Concentration icon from the More Apps screen.
2. For cuvette application (C40-Go): Select the path length depending on the used cuvette. Options are: 0.5 mm, 1 mm, 2 mm, 5 mm and 10 mm
If it is desired to heat the sample to 37°C use the toggle switch to turn on cell holder heater. When the cuvette holder has reached 37°C the toggle color changes to green. Note: Only available for cuvette applications (C40-Go). For NanoVolume application (only with N50-Go Upgrade): Select the dilution depending on the sample concentration.
Note: There is no automatic path length setting in this method. Select either a virtual dilution of 15 (path length 0.67 mm) or of 140 (path length 0.07 mm). 3. Default wavelength is 260 nm but can be changed in the range of 200900 nm (N50: 200­650 nm), depending on the sample/application. 4. Enter a factor for concentration calculation.
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6. Baseline correction is set off by default. Enabling the baseline corrections shows a list with different wavelength options: 377 nm, 604 nm, 650 nm, 770 nm (N/A N50) and 823 nm (N/A N50). Option to enter any wavelength between 200 nm and 900 nm (N50: 650 nm)
7. Option to smooth the graph with different boxcars. Options: Off, 1 = boxcar 11 (default), 2 = boxcar 21 and 3 = boxcar 61
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8. Option to set/calculate a dilution factor for manual diluted samples.
9. Apply the blank ddH20 or buffer to the illuminated sample window on pedestal for the reference measurement and select blank to initiate the reading. Note: The illumination of the sample window can be switched off in the preferences.
10. Use a lint-free laboratory wipe to clean both the sample window on pedestal and mirror in lid arm prior to applying the next sample. Note: It can be helpful to apply the blank a second time and read it as a sample to ensure a proper blank.
11. Apply sample to the sample window on pedestal and press the sample button to initiate the measurement.
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CALCULATIONS
In this method, the concentration of the sample is calculated based on the Beer-Lambert law given the user specified wavelength of interest and user defined extinction coefficient. The equations for calculating concentration without background correction are as follows:

Without background correction:

= Ð

C

Concentration (ng/µl)

Absorbance at user specified path length n (10 mm path)

Ð

Dilution factor

extinction coefficient/factor

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MORE APPS: STANDARD CURVE
Available only for C40-Go, N50-Go needs upgrade for activation of Standard Curve.
METHOD OVERVIEW
The construction of a calibration curve from multiple standards of known concentrations can be created and stored on the NanoPhotometer® Go. The standard curve can be used to quantify samples of the same type with unknown concentrations. This application provides an extremely useful tool with which to integrate, expedite and simplify the measurement and calculations involved in determining the concentration of analytes in unknown samples. If a zero concentration standard is required, include it in the number of standards to be entered using a reagent blank and entering 0.00 for concentration.
MEASUREMENT PROTOCOL
1. Select the More Apps icon from the home screen and the Standard Curve icon from the More Apps screen.
2. For cuvette application (C40-Go): Select the path length depending on the used cuvette.
If it is desired to heat the sample to 37°C use the toggle switch to turn on cell holder heater. When the cuvette holder has reached 37°C the toggle color changes to green. Note: Only available for cuvette applications (and C40-Go). 3. For NanoVolume application (only with N50-Go upgrade): Select the dilution depending on the sample concentration
Note: There is no automatic path length setting in this method. Select either a virtual dilution of 15 (path length 0.67 mm) or of 140 (path length 0.07 mm). 4. Baseline correction is set off by default. Enabling the baseline corrections shows a list with different wavelength options: 377 nm, 604 nm, 650 nm, 770 nm (N/A N50) and 823 nm (N/A N50). Option to enter any wavelength between 200 nm and 900 nm (N50: 650 nm)
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line through the origin) and 2nd order regression. 6. Select Unit
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concentrations can be deleted with Enter the concentrations of the standard curve.
8. Select Replicates none, 2 or 3
9. Once Standard curve is created or loaded it will be used for concentration calculations in the method. It might be necessary to do a blank measurement.
10. Apply sample and press the sample button to initiate the measurement. Note: Once the sample measurement is initiated it is not possible to make changes to the standard curve.
SAVING AND LOADING STANDARD CURVES
It is possible to save measured standard curves as a Stored Method. To save the standard curve push the store method button and enter a method name, select a folder and save with the Store button. Methods can be opened in the Stored Methods menu on the homescreen. Opening a saved Protein Assay method shows a message with the option to load or remeasure the standard curve.
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CALCULATIONS
Concentration is determined via the absorbance values provided by the standard curve based on the curve fit selection including the following options: linear regression, zero regression and 2nd order regression.
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CUSTOM APPS
There is an option for designing customer specific Custom Apps which can be loaded to the NanoPhotometer® Go. For more information about designing custom applications to suit individual research needs please contact Implen directly for assistance.
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STORED RESULTS
The Stored Results icon opens a directory of folders containing files of results that have been previously saved.
On the left side of the screen all available directories/storages are shown: NanoPhotometer®, Control Device, Network and/or USB flash drive (depending on availability). Pushing a storage folder shows the subfolders of this storage folder on the left side and the individual files on the right side. On the right side of the screen all saved result files of the selected folder are shown and can be opened by a long or double click. Folders can be deleted, renamed, moved or copied by pushing on the icon. It is also possible to delete, rename, move or copy files by pushing on the icon. The file path of the selected folder is shown on the top of the right file area. Note: PDF and Excel files cannot be opened on the NanoPhotometer® Go. Files need to be transferred to a computer or device where Excel or a PDF reader is installed. Note: Control device is only available on computer, tablets and smartphones not on the NanoPhotometer® Go version of the software. For data transfer via Ethernet or WiFi see page 38 Data Transfer. Backup copies are saved in the Autosave folder of the NanoPhotometer® Go (Stored Results/NanoPhotometer/Autosave) for up to ten days. After ten days the autosave files are automatically moved to an autosave archive folder. The autosave archive folder can only be accessed via NanoPhotometer® Go file server. Data in the autosave archive folder are not automatically deleted. The content of the Autosave Archive folder can be deleted via the action button in Stored Results. Make sure that a backup is created before deleting the Autosave Archive folder content.
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STORED METHODS
The stored methods icon opens the directories of folders containing methods stored by the user.
On the left side of the screen all available directories/storages are shown: NanoPhotometer®, Control Device, Network and/or USB flash drive (depending on availability). Pushing a storage folder shows the subfolders of this method folder on the left side and the individual files on the right side. On the right side of the screen all saved method files of the selected folder are shown and can be opened by a long or double click. On the top of the right area the file path of the selected folder is shown. New folders can be created by pushing on . Folders can be deleted, renamed, moved or copied by pushing on the icon. It is also possible to delete, rename, move or copy methods by pushing on the icon.
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5. PREFERENCES
System preferences can be set by selecting preferences on the home screen. The preferences menu includes: General, Dyes, Warning Messages, Network, Printer and CFR21. The preference options of the selected menu item are listed in the window on the right. Note: Preferences are not available on smartphones with screen sizes less than 7 inches.
GENERAL
Selecting General in the Preferences menu opens a window to the right of the preferences menu with the following options: Date and Time, Display, About, Storage and Illumination Sample Window (N50-Go).
DATE AND TIME
Within the Date and Time it is possible to set the actual date and time of the NanoPhotometer® Go or change the time zone.
To change the time zone or the date and/or time, push on the appropriate field to open the selection options. The changed settings are shown below the time zone field. To apply the changes, the NanoPhotometer® Go must be rebooted. Start the reboot by pushing on the Set and Reboot button.
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DISPLAY
Brightness: adjustment of the built-in screen brightness
ABOUT
In About the following information of the NanoPhotometer® Go are shown: NanoPhotometer® Version, Serial Number, Ethernet IP Address, WiFi IP Address, Hardware Version, Firmware Version, Time & Date of Initialization Test and Status of Initialization.
STORAGE
Shows the total storage capacity and the free space of the internal NanoPhotometer® Go storage.
ILLUMINATION SAMPLE WINDOW
Toggle switch to switch on/off the illumination of the sample window (for N50-Go only)
DYES
There is a list of preprogrammed dye-labels for both nucleic acid dyes and protein dyes. To toggle between the nucleic acid and protein list push on the Nucleic Acid/Protein buttons in the header.
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Each dye has either a lock icon ( ) in front of the dye name indicating that the dye is locked and cannot be changed or a delete symbol ( ). The delete option is only available for unlocked and not preprogrammed dyes. Selecting a dye name opens a new screen with the dye information: dye name, absorbance maximum dye (nm), dye-dependent extinction coefficient dye (M-1 cm-1), and dyedependent correction factor as well as the option to show the dye in the parameter list of the application (Nucleic Acid or Protein UV).
Note: It is not possible to delete a dye from the default factory list; custom dyes can be deleted if they are not locked. It is possible to add a new dye to the list by selecting the + button to add a new dye. A window will open where it is possible to enter the: dye name, dye absorbance maximum (nm), dye- dependent extinction coefficient dye (M-1
cm-1), and dye-dependent correction factor. There is a toggle switch available to lock the dye to prevent deleting a dye from the dye list accidentally.
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WARNING MESSAGES
BLANK CONTROL
Toggle switch to switch on/off the Blank ControlTM of the NanoPhotometer® Go. Note: Blank ControlTM is available for all NanoVolume methods (N50-Go).
SAMPLE QUALITY CONTROL
It is possible to change the upper and the lower limit of the ratio alert warning messages. Default values for nucleic acid ratios are: 260/230 ratio 1.8 A – 3 A and 260/280 ratio 1.65 A – 2.5 A. Default value for Protein UV ratio is: 260/280 ratio is 0.7 A.
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NETWORK
Selecting Network in the Preferences menu opens a window to the right of the preferences menu with the following options: Network Settings, WLAN Settings, File Server Access and Network Folder.

References

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