AOAC RI ERP Final Action Recommendations eBook

Collaborative Study Approved Protocol Expert Review Panel Use Only

(Applicable to the detection of Salmonella in raw ground beef (25 g and 375 g), chicken carcass rinse, 1 hot dogs, beef trim, ground beef with soy, ground turkey, cream cheese, fresh bagged lettuce, orange 2 juice, frozen peas, peanut butter, shell eggs, shrimp, white pepper, ice cream, non-fat dry milk, dried 3 eggs, cocoa, and infant formula, and dry pet food (375 g), and on stainless steel, ceramic tile, and plastic 4 surfaces) 7 Kits — Reagents used in the BAX® System should pose no hazards when used as directed. Dispose of 8 lysate, PCR mixture and other waste according to your site practices. 9 Cycler/detector — Only qualified laboratory personnel should operate the cycler/detector. Do not 10 attempt to repair the instrument. Live power may still be available inside the unit even when a fuse has 11 blown or been removed. Refer to the BAX® System User Guide for maintenance procedures when 12 cleaning the unit or changing a fuse. The heating block can become hot enough during normal operation 13 to cause burns or cause liquids to boil. Wear safety glasses or other eye protection at all times during 14 operation. 15 Enrichment Broths — All enrichment broths may contain varying pathogens whether they contain 16 Salmonella or not and thus should be sterilized and disposed of using proper procedures following 17 any culture-based confirmatory steps. 22 bacterial DNA, which is stable and unaffected by growth environment. The BAX® System simplifies 23 the PCR process by combining the requisite primers, polymerase and nucleotides into a stable, dry, 24 manufactured tablet already packaged inside the PCR tubes. After amplification, these tubes 25 remain sealed thus significantly reducing the potential for contamination with one or more 26 molecules of amplified PCR product in future tests. 27 This automated BAX® System method uses fluorescent detection to analyze PCR product. One 28 PCR primer for each target (one Salmonella- specific target and an internal control) contains a 29 fluorescent dye (two different dyes, one for each target) as a constituent of the primer as well as a 30 quencher (the uni-molecular combination of a primer, fluorescent dye and quencher constitute a 31 Scorpion™ Probe). When not incorporated into a PCR product, the Scorpion™ Probe has a hair-pin 32 loop structure which keeps the dye and quencher in close proximity. When incorporated into a PCR 33 product, the dye and quencher are spatially separated due to an internal hybridization, which 34 causes an increase in emission signal. The BAX® System measures the magnitude and 35 characteristics of fluorescent signal change. An analysis by the BAX® System software algorithm 36 then evaluates that data to determine a positive or negative result. 5 6 Caution: 18 19 20 21 A. Principle The BAX® System uses the Polymerase Chain Reaction (PCR) to amplify a specific fragment of

37 38 39 40 41 42

B. Apparatus

(a) BAX® System Q7 cycler/detector with computer workstation.

(b) BAX® System application software.

9