AOAC-RI ERP Book Micro Jan 19 2017

OMAMAN-35 A : Collaborative Study Manuscript For ERP Use Only January 2017

In the rare event of any unusual light output, the algorithm labels this as “Inspect.” 3M recommends the user to 1 repeat the assay for any Inspect samples. If the result continues to be Inspect, proceed to confirmation test using 2 your preferred method or as specified by local regulations. 3 4 In the event of discordant results (presumptive positive with the 3M Molecular Detection Assay 2 - E. coli O157 5 (including H7), non-confirmed by one of the means described above, and in particular for the latex agglutination 6 test), the laboratory must follow the necessary steps to ensure the validity of the results obtained. 7 8 If you have questions about specific applications or procedures, please visit our website at 9 www.3M.com/foodsafety or contact your local 3M representative or distributor. 0 1 Appendix A. Protocol Interruption: Storage and re-testing of heat-treated lysates 2 1. To store a heat-treated lysate, re-cap the lysis tube with a clean cap (see “LYSIS”, 4.5) 3 2. Store at 4 to 8°C for up to 72 hours. 4 3. Prepare a stored sample for amplification by inverting 2-3 times to mix. 5 4. Decap the tubes. 6 5. Place the mixed lysate tubes on 3M Molecular Detection Heat Block Insert and heat at 100 ±1°C for 5 ±1 7 6. Remove the rack of LS tubes from the heating block and allow to cool in the 3M Molecular Detection Chill 9 Block Insert at least 5 minutes and a maximum of 10 minutes. 0 7. Continue the protocol at the ‘Amplification’ section detailed above. 3 4 For this qualitative, unpaired collaborative study, the 3M Molecular Detection Assay (MDA) 2 – E. coli O157 5 (including H7) method was compared to the USDA FSIS MLG 5.09 reference method for raw ground beef (325 6 g). A total of 15 laboratories throughout the United Statesparticipated in this study, with 10 laboratories 7 submitting data for the raw ground beef. See Table 1 for a summary of laboratory participation. Each laboratory 8 analyzed 36 test portions for each method: 12 inoculated with a high level of E. coli O157:H7, 12 inoculated with a 9 low level of E. coli O157:H7, and 12 un-inoculated controls. 0 A background screen of the matrix indicated an absence of indigenous E. coli O157:H7 in the ground beef.Due to 1 the amount of raw ground beef needed for the evaluation (~900lbs.), ten (10) replicate 325 g test portionswere 2 screened for E. coli O157:H7. All test portions produced negative results for the target analyte. 3 The individual laboratory and sample results are presented in Table1 of the Supplementary Materials. 4 Table2016.1A summarizes the inter-laboratory results forthe ground beef, including POD statistical analysis [9]. 5 As per criteria outlined in Appendix J of the AOAC Validation Guidelines, fractional positive results were 6 obtained.For each matrix, the level of E. coli O157:H7was determined by MPN on the day of initiation of analysis 7 by the coordinating laboratory. MPN results are presented in Tables 2016.2A.The APCresults for each 8 collaborating laboratory are presented in Table 2of the Supplementary Materials. 1 2 Raw ground beef test portions were inoculated at a low and high level and were analyzed (Table 1 of the 3 Supplementary Materials) for the detection of E. coli O157:H7. Un-inoculated controls were included in each 4 analysis. Laboratory 3 was unable to confirm their samples using the USDA method, Laboratory 7 5 reportedcontamination of the E buffer and was not able to return to the enrichments to re-confirm, Laboratory 10, 6 13, and 15 did not follow the alternative method such that results could not be used. All other laboratories 7 submitted a full set of data for each method. The MPN levels obtained for this test portion, with 95% confidence 8 AOAC Research Institute Expert Review P nel Use Only minutes. 8 1 2 Results of Collaborative Study 9 0 Raw Ground Beef (73% Lean) (325 g Test Portions)

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