AOAC-RI ERP Micro June 2016

OMAMAN-29 D/ PTM Validation Report 111501 OMA ERP - June 2016 ERP Use Only

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Results were identical after 24 and 26 hours of sample enrichment. For the low inoculation level of the MDA 2 - Listeria assay, there were 15 presumptive positives and 15 confirmed positives following the USDA/FSIS MLG 8.09 reference method confirmation procedure. There were 11 observed positives for the reference method. For the high inoculation level, there were 5 presumptive positives and 5 confirmed positives following the USDA/FSIS MLG 8.09 reference method confirmation procedure. There were 5 confirmed positives following the reference method. Detailed results of the POD analyses are presented in Tables 9 and 13.

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Product Consistency (Lot-to-Lot) and Stability Studies

Methodology

Three lots of MDA 2 - Listeria test kits, 1 newly manufactured, 1 at the middle of its expiration, and 1 at or slightly beyond expiration were analyzed to determine the stability of the assay. For the MDA 2 - Listeria test kit, one Listeria monocytogenes ATCC 7644 isolate was analyzed at a fractional positive level (2-8 positives), and testing 10 replicates. One non- Listeria isolate, Enterococcus faecalis ATCC 29212, was analyzed at the growth level achieved in a non-

selective broth, testing 5 replicates.

Results

For the lot-to-lot/stability evaluation of the MDA 2 - Listeria assay, there were 6 presumptive positives out of 10 replicates for all three lots at the low inoculation level. For the 5 uninoculated control test portions, there were 0 presumptive positives out of 5 replicates for all three lots. The five control test portions were negative. See Table 16 for lot-to-lot/stability results.

Robustness Study

Methodology

The robustness evaluated the ability of the method to remain unaffected by minor variations in method parameters that might be expected to occur when the method is performed by an end user. Three different parameters were evaluated, which are presented in Table 18. For each parameter, one Listeria monocytogenes ATCC 7644 isolate was analyzed at a fractional positive level (2-8 positives), testing 10 replicates. One non- Listeria isolate, Enterococcus faecalis ATCC 29212, was analyzed at the growth level achieved in a non-selective broth, testing 5 replicates.

AOAC Research Institute Expert Review Panel Use Only

Table B: Robustness Parameters

Parameter Lysis Time

Low Value 13 minutes

Nominal Value

High Value 17 minutes

15 minutes

Enrichment Volume

18 µL 18 µL

20 µL 20 µL

22 µL 22 µL

Lysate Volume

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Table C: Robustness Parameter Variations

Treatment

Lysis Time

Volume

Volume

# of Replicates