BAX® System Q7 Ready Reference for Yeast and Mold PCR Assay
A concise guide for performing Yeast and Mold PCR Assays using the Hygiena BAX System Q7.
Procedure Overview
Step 1: Homogenize Sample
Homogenize sample in a 1:10 dilution according to the food type. Refer to the User Guide or table on the back of this reference card for specific instructions based on food type.
Step 2: Determine Sample Volume
Determine the sample volume to be tested.
Step 3: Transfer Sample
Transfer the sample to a disrupter tube. The pooled sample protocol requires triplicate disrupter tubes.
Step 4: Incubate Disrupter Tubes
Incubate disrupter tubes at 25°C for 44 hours.
Step 5: Add DNA Stabilizer
Add 20 µL of DNA stabilizer to the disrupter tubes.
Step 6: Agitate
Agitate samples in the disrupter device.
Step 7: Create Rack File
Create a rack file for the assay.
Step 8: Add Protease
Add 150 µL of protease to the YM lysis buffer (12 mL). The lysis reagent mixture can be stored at 2-8°C for up to one week.
Step 9: Transfer Lysis Reagent
Transfer the lysis reagent prepared in step 8 to cluster tubes (200 µL per tube).
Step 10: Transfer Disrupted Samples
Transfer disrupted samples to cluster tubes (20 µL per tube). For the pooled sample protocol, pool volumes from disrupter tubes into 1 cluster tube.
Step 11: Heat Cluster Tubes
Heat cluster tubes for 0:20 at 95°C. Alternatively, steps 11 and 12 can be performed using the Hygiena™ Automated Thermal Block. Refer to the Automated Thermal Block User Guide for details.
Step 12: Cool Cluster Tubes
Cool cluster tubes in a cooling block for 0:10 at 37°C, followed by 0:05.
Step 13: Initialize Cycler
Initialize the cycler. The system will indicate when it is ready for loading.
Step 14: Arrange PCR Tubes
Arrange PCR tubes in the cooling block.
Step 15: Hydrate PCR Tablets
Hydrate PCR tablets with 50 µL of lysate from step 12.
Step 16: Run Program
Place tubes in the cycler and run the program.
Step 17: Review Results
Unload samples and review results on screen. Refer to the User Guide for detailed interpretation.
Result Indicators:
- Negative: ✔️
- Positive: ➕
- Indeterminate: ❓
- Signal error: ❓
Sample Protocols
Pooled Sample Protocol
This ultra-sensitive protocol uses pooled samples from three disrupter tube enrichment replicates for action levels of 10-50 cfu/g.
| If your action level is: | Then transfer this volume of homogenate to 3 disrupter tubes: | And pool these volumes of disrupted sample for testing: |
|---|---|---|
| 10 cfu/g | 400 µL | 7 µL from 3 replicates |
| 20 cfu/g | 200 µL | 7 µL from 3 replicates |
| 50 cfu/g | 80 µL | 7 µL from 3 replicates |
Non-Pooled Sample Protocol
This protocol for yeast and mold testing can be used without pooling for action levels of 25 cfu/g or above.
| If your action level is: | Then use this volume of homogenate: |
|---|---|
| 25 cfu/g | 400 µL |
| 50 cfu/g | 200 µL |
| 100 cfu/g | 100 µL |
| 500 cfu/g | 20 µL |
| 1000 cfu/g | 10 µL |
