hygiena KIT230103 foodproof Staphylococcus Detection LyoKit User Guide
- May 15, 2024
- Hygiena
Table of Contents
hygiena KIT230103 foodproof Staphylococcus Detection LyoKit
PCR kit for the qualitative detection of the most important coagulase-positive Staphylococcus species including the simultaneous identification of Staphylococcus aureus.
PROGRAM SETUP
Program your real-time PCR instrument before setting up the PCR reactions. Select the following channels:
- FAM (coagulase-positive Staphylococcus spp.), HEX (Staphylococcus aureus) and ROX (Internal Control).
Pre-incubation: 1 cycle
Step 1: 37 °C for 4 min
Step 2: 95 °C for 5 min
Amplifi cation: 40 cycles
Step 1 : 95 °C for 5 sec
Step 2*: 60 °C for 60 sec
- Fluorescence detection
For some real-time PCR instruments the probe quencher as well as the usage of
a passive reference dye has to be specified. This kit contains probes with a
non-fluorescent “dark” quencher and no passive reference dye.
A Color Compensation is necessary for users of the LightCycler® 480 System:
Color Compensation Set 5 (Product No. KIT230011).
For the Dualo 32® R2 real-time PCR instrument, please open the software, click on ‘New’, and select the respective template file. Template files can be added by clicking on ‘Add’ in the ‘Select template file’ window.
DATA INTERPRETATION
Verify results of positive (Control Template) and negative controls (H2 O),
before interpreting sample results.
Always compare samples to positive and negative control. Review data from each
channel and interpret results as described in the table.
| FAM | HEX | ROX | Result Interpretation |
|---|---|---|---|
| + | + | + or – | Positive for S. aureus |
| + | – | + or – | Positive for coagulase-positive Staphylococcus spp. other than |
S. aureus
–| –| +| Negative for coagulase-positive Staphylococcus spp.
–| –| –| Invalid
If the Cq value for FAM or HEX is > 35, the signal may have been caused by DNA
of dead bacteria.
In this case, a prolongation of enrichment and a new analysis is recommended.
PREPARATION OF THE PCR MIX
Take appropriate precautions to prevent contamination, e.g., by using filter tips and wearing gloves.
-
PLACE STRIPS IN RACK
Take needed number of PCR tube strips out of aluminum bag.
Important: close bag tightly afterwards. Place strips in a suitable PCR tube rack.
If needed, gently tap the tubes to move the lyophilized pellets to the bottom of all tubes. -
DECAP
Carefully open strips immediately before filling and discard caps.
Do not leave open longer than necessary. -
ADD SAMPLES AND CONTROLS
Pipette 5 to 25 µL of samples, 25 µL negative control (colorless cap) or Control Template (purple cap) into respective wells.
Varying sample volumes are due to different enrichment broths and DNA extraction procedures. If using less volume, add PCR-grade H2 O to reach a total volume of 25 µL. -
SEAL
Carefully seal the tubes with the provided 8-cap strips. -
MIX
Resuspend pellet after sealing by mixing thoroughly.
Alternatively, resuspend pellet by pipetting up and down multiple times in Step 3. -
CENTRIFUGE
Briefly spin strips, e.g., 5 seconds at 500 – 1,000 x g, in a suitable centrifuge. -
START REAL-TIME PCR RUN
Cycle samples as described above.
Place tubes in a vertical, balanced order into the cycler, e.g., two strips can be placed in the first and last column.
CUSTOMER SUPPORT
foodproof®
Staphylococcus
Detection LyoKit
KIT230103 /04 /05
Kit for 96 reactions
Store kit at 2 to 8 °C
For food testing purposes
FOR IN VITRO USE ONLY
Made in Germany
Hygiena® | Camarillo, CA 93012 USA |
diagnostics.support@hygiena.com |
www.hygiena.com