AOAC-RI ERP Micro June 2016

OMAMAN-29/OMAMAN-30 Combined Collaborative Study Protocol OMA ERP - June 2016 ERP Use Only

3M MDA 2 Listeria species and L. monocytogenes Collaborative Study OMA-2016-MONTH-XXX

4.6.1.2 Raw chicken breast - aseptically add the 25g test portion to 475 mL Demi Fraser broth. 4.6.1.2.1 Homogenize thoroughly in a filter bag for 2 ± 0.2 min 4.6.1.2.2 Incubate 28-32 hr at 37 ± 1°C. 4.6.2 Prepare the 3M Molecular Detection speed loader tray, chill block insert, heat block insert and instrument. Equilibrate the LS tubes to room temperature (20-25 ºC) by setting LS tube overnight 16-18 hours. Or on the laboratory bench for at least 2 hours. 4.6.3 Invert the capped LS tubes to mix, up to 4 hours before use. 4.6.4 Transfer 20 µL enriched sample into a LS tube. 4.6.5 Transfer 20 µL Negative Control [NC], sterile enrichment medium (e.g. Demi-Fraser Broth) into a Lysis Solution [LS] tube after all enriched samples have been completed. Do not use water as a NC. 4.6.6 Place uncovered LS tubes in 3M Molecular detection Heat block, heat 15±1 min at 100±1°C. LS Solution will change from pink (cool) to yellow (hot). 4.6.7 Place LS tubes (without rack lid) in chill block insert (at ambient temperature) for 5-10 min. Lysis solution in LS tube will revert to pink color. 4.6.8 Transfer 20 µL sample lysate from the upper portion of fluid in the LS tube into regent tube. Mix gently by pipetting up and down 5 times. 4.6.9 Transfer 20 µL of NC lysate into a reagent tube. 4.6.10 Transfer 20 µL of NC lysate into a Reagent Control (RC) tube. 4.6.11 When all samples have been transferred, load capped tubes into speed loader tray and close lid. 4.6.12 Start assay. 4.6.13 Results and Interpretation - An algorithm interprets the light output curve resulting from the detection of the nucleic acid amplification. Results are analyzed automatically by the software and are color-coded based on the result. A Positive or Negative result is determined by analysis of a number of unique curve parameters. Presumptive positive results are reported in real-time while Negative and Inspect results will be displayed after the run is completed. Presumptive positive results should be confirmed using your preferred method or as specified by local regulations. NOTE: Even a negative sample will not give a zero reading as the 3M MDS and 3M MDA amplification reagents have a “background” relative light unit (RLU). In the rare event of any unusual light output, the algorithm labels this as “Inspect.” 3M recommends the user to repeat the assay for any Inspect samples. If the result continues to be Inspect, proceed to confirmation test using your preferred method or as specified by local regulations. AOAC Research Institute Exp rt Review Panel Use Only

5/4/16