Jack Rowe
Certified Laboratories, Inc.
Bridging Science with Service
Since 1926
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AOAC Performance Tested Method (RI)
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AOAC Official Methods of Analysis
Rapid increase in Proprietary Methods
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Speed to market critical
AOAC OMA
‣ Higher Cost, More Time
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Performance Tested Methods
‣ Kicked off in 1991
‣ Opportunity to achieve some level of validation in less
time
‣ Often serves as a “pre-collaborative” until full OMA is
completed
Matrix Study
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One laboratory
1-4 Foods or Surface
Inclusivity, Exclusivity,
Robustness, Stability and Lot
to Lot Variation
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Pre-Collaborative Study and
Collaborative Study
Harmonization Collaborative Study
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Selection of matrices associated with
pathogen
Natural vs. Artificial contamination
Strain types
Competing flora
Fractional results
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Quantitative
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8 laboratories minimum, 1-6 matrices
4 inoculation levels/matrix
2 test portions per level per matrix per method
Qualitative
• 10 laboratories minimum, 1-6 matrices,
• 3 inoculation levels per matrix,
• 6 test portions per level per matrix per method
• Method compared to Reference Method
FDA BAM
 USDA FSIS
 AOAC
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Official Methods of Analysis
2011.03
Salmonella in a Variety of Food
Validated Matrices
Foods/Eggs and Egg Products,
Eggs and Egg Products/Liquid Eggs,
Eggs and Egg Products/Eggs,
Dairy Products/Ice Cream,
Vegetables/Spinach,
:
Spinach/Fresh Spinach,
Fish/Shrimp,
Nuts and Nut Products/Peanut Butter,
Peanuts/Peanut Butter,
Meat and Meat Products/Turkey,
Meat and Meat Products/Pork,
Meat and Meat Products/Roast Beef,
Meat and Meat Products/Pork Sausage,
Meat and Meat Products/Chicken,
Dairy Products/Milk,
Milk/Whole Milk,
Foods/Fish,
Nuts and Nut Products/Pecans,
Baked Goods/Cake Mix,
Grains/Dry Pasta,
Spices and Condiments/Black Pepper,
Milk/Dried Milk,
Eggs and Egg Products/Dried Egg Yolks,
Cacao Bean and Its Products/Chocolate,
Foods/Fruits and Fruit Products,
Fruit Juices/Orange Juice,
Pet Foods,
Water
Domestic Imports
U.S. food import volume, by food group
Food
group
1999
Volume of U.S. food imports 1/
Live meat animals (1000) 2/
Meats
Fish and shellfish
Dairy
Vegetables
Fruits 3/
Nuts
Coffee and tea
Cereals and bakery
Vegetable oils
Sugar and candy
Cocoa and chocolate
Other edible products
Beverages (1000 KL) 4/
6,058.4
1,439.0
1,677.8
242.5
4,767.5
8,193.8
272.0
1,709.4
6,687.6
2,320.8
322.1
923.7
705.9
2,854.9
2012
7,908.0
1,335.8
2,354.2
209.8
8,545.0
11,569.6
443.0
2,093.8
10,184.4
5,178.1
3,730.6
1,237.4
1,447.3
5,820.1
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FDA will accept all matrices validated by the
manufacturer for the AOAC study
Other matrices must be verified by laboratory
to determine if the method is suitable for the
matrix
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Amaranth
Coconut
Cumin
Egg Cracklet
Hing Powder
Tahina/Sesame Seeds
Octopus
Spike level must not exceed 30 cells
(25 gram sample)
Seven successful validations must occur
(No false negatives or positives)
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Salmonella culture
Atypical or Typical
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Serial dilution in Phosphate Buffer
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Plated in Duplicate 1ml and 0.1ml
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Selective Agar
Hektoen or XLD
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Count plates after incubation per BAM
Example
Dilution 4
Dilution 4
125
113
Dilution 5
Dilution 5
125 + 113 / 2 = 119 cfu/ml
14
12
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Dilution 4 tube contains 119 cfu/ml
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X ml x 119 cfu/ml = 30 cfu
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30 cfu/119 cfu/ml = X ml
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X = 0.252 or 250 ul
Prepare sample per BAM (or BioMerieux)
instructions
Weigh out 25 gram sample and spike with
calculated volume
Plate the calculated spike volume in
duplicate on selective agar
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Incubate per normal protocol
Confirm counts from spike volume do not
exceed 30 cells
Control sample must produce a negative
result. If the sample is “presumptive positive”
it contains an unspecified amount of
Salmonella and demonstration of < 30 cfu/
25 gram sample is not possible
Abandon Verification
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All controls must give appropriate reactions
Positive Cultural Control
Negative Cultural Control
Diluent Blank
The Presumptive Positive spike must be
confirmed following reference method
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Biochemical Identification of Spike, Negative
and Positive Control
Media preparation logs for all media used
Autoclave Sterilization Records for all media
Media Sterility records for all media used
Balance Calibration Check Records
Temperature Records for all Appliances
Water Quality Records
Confirmation serves as Document of
Productivity