The AOAC International

Rapid Methods
Validation Process
Scott Coates Zerlinde B. Johnson
Managing Director
Manager, Technical Programs
AOAC Research Institute
AOAC Research Institute
Overview of Presentation
 AOAC International
 AOAC Research Institute
 Consensus Programs
 Validation Programs
 Publications & Documentation of Approved
Methods
 Survey of PTM & OMA approved methods
 AOAC INTERNATIONAL
• Non-profit organization with international membership
• Consensus organization
• Established in 1874 to review, validate & publish analytical methods
• 125th Anniversary in 2009!
• best known in food and agriculture
• chemistry and microbiology methods for Official Methods of
AnalysisSM status (OMA) “Big Red Book of Methods”
• OMA methods are recognized in the U.S. Code of Federal
Regulations and are legally defensible in court
worldwide
• staff of approx. 40 in Gaithersburg, Maryland, USA
 AOAC Research Institute
• Incorporated in December 1991 as a subsidiary of
AOAC INTERNATIONAL
• Non-profit, international, scientific organization
• Administers the Performance Tested MethodSM Program
• Chemistry and microbiology methods
• Provides rapid entry point into the AOAC validation process and
provides a rapid approval process
• Provides an independent third-party review of test kit
performance claims
• Staff with industry experience
AOAC Consensus Programs
 Helping solve scientific-based problems with
scientific-based solutions
 AOAC brings together people from industry,
government, public health, academia and end users
to discuss health, business and trade issues
 Setting of acceptance criteria for methods for
particular target analyte (B. anthracis, F. tularensis,
Y. pestis, drug residues in commodities to name a
few issues)
AOAC Validation Programs
 AOAC International

 Official Methods of AnalysisSM (OMA)

 AOAC Research Institute

 Performance Tested MethodsSM

(PTM)
 Harmonized Program

 PTM-OMA
Method Validation Guidelines
For Microbiology:
 AOAC Guidelines For Validation Of Qualitative And Quantitative
Food Microbiological Official Methods Of Analysis published J.
Assoc. Off. Anal. Chem. 85, 1187-1203 (2002)
 ISO 16140: Protocol for the validation of alternative methods

For Chemistry:
 "Guidelines for Collaborative Study of Procedure to Validate
Characteristics of a Method of Analysis," J. Assoc. Off. Anal.
Chem. 72, 694-704(1989)
 Used by ISO, AOAC, NMKL, and many others

Used when applicable by both AOAC validation programs

 Official Methods of
Analysis
 Two Phase Validation: Pre-Collaborative and Collaborative
 Number of Collaborative Labs Required:
 Quantitative method: 8 laboratories minimum
 Qualitative method: 10 laboratories minimum

 Data Review by Appropriate Methods Committee and

General Referee
 Validation Time: 12-18 Months Minimum to First Action
 Final Action can be obtained two years after First Action
 Official Methods of
Analysis
Validation Study Design
Pre-Collaborative Studies Collaborative Study
 Inclusivity  Method Comparison
 Exclusivity  8-10 labs
 6 foods
 Method Comparison
 USDA, FDA, ISO or
 15-20 foods
other appropriate
 USDA, FDA, ISO or other
reference methods
appropriate reference
may be used
methods may be used
 Method Comparison Study
 Determines matrices that can be claimed to be validated
 A method can NOT be validated for all foods Qualitative methods:
 Pre-Collaborative Study and Performance Tested Method studies require 20
replicates at each inoculation level and 5 uninoculated controls or 20 replicates
of naturally contaminated
 Collaborative Study (Ring Trial) requires 6 replicates at each inoculation level
and 6 uninoculated controls per laboratory
 MUST obtain fractionally positive results

 Quantitative methods require five replicates at each of three inoculated levels

and five uninoculated controls or 3 lots of naturally contaminated foods
 For OMA approval, the candidate method must perform equal to or better than
the reference method
 Performance
Tested MethodsSM
 Evaluation of the test kit performance claims
 Two-part Validation – Internal Studies and Independent Study
 One Independent Laboratory required – contracted by AOAC RI
 Data review by two Expert Reviewers and General Referee
 Companies licensed to use certification mark in advertisements and
packaging for one year increments – annual reviews
 Validation Time: can be less than 6 months
 List of Performance Tested Methods published on the internet at
http://www.aoac.org/testkits/testedmethods.html
 Consulting service available for drafting of the validation study
 Performance
Tested MethodsSM
 Test Kit Claims
 Variety of Foods
 Requires testing of 2 foods per food groups; total of 10 foods
 Groups of Foods
 Requires testing several foods from the particular food group
 Select Foods
 Requires testing of all select foods
 Individual Foods
 One matrix claim
 Environmental Surfaces
 9 surfaces
 Pure / Mixed Cultures
 Performance
Tested MethodsSM
 Food Categories  Environmental Surfaces
 Meat products  Stainless steel
 Poultry products  Food-grade painted
 Fish & seafood products surfaces
 Dairy products  Ceramic, glass or other
 Chocolate & bakery earthen-glazed material
products  Cast iron
 Animal feeds  Plastic
 Pasta  Rubber
 Miscellaneous (dressings,  Sealed concrete
spices, mayonnaise,  Wood
eggs/egg derivates, cereals  Air filter material
and rice)
 Performance
Tested MethodsSM
Validation Study Design
Internal Study
 Conducted by the test kit manufacturer
 Inclusivity Panel (30-50 strains; 100 strains for Salmonella spp.)
 Exclusivity
 Method Comparison
 choice of reference methods
 FDA, USDA, ISO or other appropriate reference method
 Ruggedness
 Stability
 Lot-to-Lot Variation
 Performance
Tested MethodsSM
Independent Study
 Conducted in a lab contracted by AOACRI
 Intended to verify the internal study data
 Subset of internal study work
 Method Comparison
 1 laboratory
 1-2 foods
Performance Tested MethodsSM

Independent Laboratories
 Several laboratories to  Contracted with AOAC
choose from. RI, the lab performs
 Each laboratory is evaluation based upon
screened and trained protocol and kit
 Submits a bid for the package insert.
study based upon  Report submitted to
approved testing Research Institute
protocol
Performance Tested MethodsSM
Expert Reviewers
 AOAC General Referee  Two Expert
Reviewers
 Approves all testing
protocols and reviews  50% of Expert
the final Validation Reviewers are from
Report outside of the US

 Reviews Validation
Report
Performance Tested MethodsSM
Review Process
 Test Kit Manufacturer  Reviewers
 Compiles validation  Review validation report
report of internal report for technical and editorial
and independent criteria and package
laboratory information insert
 Provide commentary on
 Reply to reviewers’ report and/or insert
comments and/or  Performance TestedSM
suggestions status granted upon
consensus
Performance Tested MethodsSM
Certification
 Test kit manufacturers review
& respond to all certification
documentation
 Test kit is listed on the AOAC
 License agreement and use of RI validated methods website
the certification mark
 Certificate and study report
 Article in AOAC Magazine made available on the AOAC
Inside Laboratory validated methods website for
Management (ILM) users to download

 Publication of study in Journal  Annual renewal to ensure test

of AOAC INTERNATIONAL kit has not been changed since
validated
Performance Tested MethodsSM
Certificate
Harmonized Program
 Administered through AOAC Research Institute
 PTM studies are equivalent to the OMA
Pre-Collaborative Study
 Obtain Certification Mark first
 Continue on to Collaborative Study to obtain
OMA First Action
Harmonization Overview

4) Official Methods of Analysis

3) OMA Collaborative Study
2) Performance Tested Methods Study
1) AOAC Research Institute Consulting

OUTCOMES:
• 2 AOAC approvals (PTM & OMA)
• 2 published studies in Journal of AOAC Int’l
• Use of Certification Mark
Performance Indicators
 Inclusivity rate  Reproducibility
 Exclusivity rate  Method Agreement or
 Sensitivity rate McNemar’s Chi-Square

 Specificity rate  Limit of Detection or Limit

 False Positive rate of Quantitation

 False Negative rate  Relative Standard

Deviation
 Repeatability
 Method Publications
• Official Methods of AnalysisSM  Performance Tested
MethodsSM
• Method published in Official
 Published in Journal of AOAC
Methods of Analysis of AOAC
INTERNATIONAL
INTERNATIONAL (OMA)

 Article published in the AOAC

• Published in Journal of AOAC
magazine Inside Laboratory
INTERNATIONAL
Management

• Article published in the AOAC

 Published on the AOAC
magazine Inside Laboratory
validated methods website
Management (ILM)
List of Approved
Performance Tested MethodsSM
Survey of
Performance Tested MethodsSM
 Survey is not all inclusive
 Intended to provide a snap-shot of approved kits
 Approved methods come and go.
 Visit the AOACRI website to find out if a kit is
PTM approved
 http://www.aoac.org/testkits/testedmethods.html
Applied Biosystems

 TaqMan® Salmonella
enterica Detection Kit
BioControl Systems, Inc

 SimPlate for TPC (Total Plate

Count)
 Assurance GDS™ for
Salmonella,
 Assurance GDS™ for Listeria
spp.
 Assurance GDS™ for L.
monocytogenes
 Assurance EIA™ for Listeria
 VIP® Listeria
BD Diagnostics
 BBLTM CHROMagarTM
 Salmonella
 Listeria
 O157
 Staph aureus
bioMerieux, sa
 VITEK® 2 GP
(Gram Positive
Identification Card)

 VITEK® 2 GN
(Gram-Negative
Identification Card)

 VITEK® 2 BCL
(Bacillus Identification
Card)
bioMerieux, sa
 VIDAS® Listeria (LIS) Assay

 VIDAS® E. coli O157 (ECO) Test with O157:H7 ID Agar: 8 Hour and
24 Hour Methods

 VIDAS® Staph enterotoxin II (SET2)

 VIDAS® Listeria species Xpress (LSX) with Ottaviani Agosti Agar

(OAA)

 VIDAS Listeria DUO (LDUO) Ottaviani Agar

bioMerieux, sa
 TEMPO®
 TVC (Total Viable Count) Method
 EC ( E. coli ) Test
 CC (Coliform Count)
 EB (Enterobacteriaceae)
Bio-Rad Laboratories
 RAPID’ L.Mono
 ‘ Salmonella Agar
 ‘ E.coli O157:H7 Agar
 ’ E. coli 2 Agar
 ' Staph Agar
 ' Listeria spp. Agar
Bio-Rad Laboratories
 iQ-Check Listeria spp. Real-Time PCR Test Kit
 iQ-CheckTM Salmonella II Real-Time PCR Test Kit
 iQ-CheckTM E. coli O157:H7 II Real-Time PCR Test Kit
Chisso Corporation

 Sanita-kun Total Plate Count

 Sanita-kun Coliforms
Centrus International
/ FOSS A/S
 Soleris/
MicroFoss Coliform Test
Daiken Industries, Ltd.
 DOX-60F / 30F
TVC Kit
Dupont Qualicon, Inc.
 BAX® System with Automated Detection PCR Assay for:
 Staphylococcus aureus
 Screening L. monocytogenes
 Screening Listeria genus
 Screening for Salmonella
 E. coli O157:H7 Kit
 Screening E. coli O157:H7 MP
 Campylobacter jejuni / coli / lari
 BAX® System with Automated Detection PCR Assay &
Collection Kit for:
 Listeria spp.
Dupont Qualicon, Inc.
 DuPont Lateral Flow System
 Listeria Test Kit
 E. coli O157 Test Kit
 Salmonella Test Kit
Idaho Technology, Inc.

 R.A.P.I.D® LT Food
Security System for
Salmonella Detection
Matrix MicroScience Ltd
 PATHATRIX
 Salmonella species Pooling Test System
 Listeria species Pooling Test System
 Listeria species Test System
 Listeria/Salmonella species
 Salmonella species
 E.coli O157 Test System
 Combined E.coli and Total Coliform Test
Merck KGaA
 Singlepath®
 E. coli O157 Lateral Flow
Assay
 Salmonella Lateral Flow
Assay
 Campylobacter Lateral
Flow Assay
 Duopath® Verotoxin
Lateral Flow Assay
Neogen Corporation
 Reveal
 Salmonella spp.
 Listeria Test System

 GENE TRAK
 Salmonella DLP Assay
 Listeria DLP Assay
Neogen Corporation
 GeneQuence
 Salmonella Microwell assay
 Listeria Microwell Test
 Listeria monocytogenes Test
 BetaStar®
US beta-lactam Assay Test
Nissui
Pharmaceutical Co.

 Compact Dry
 Total Count
 Compact Dry YM
 Compact Dry CF
 Compact Dry EC
Strategic
Diagnostics Inc.
 RapidCheck for:
 E. coli O157 (including H7) Lateral Flow Assay
 Salmonella Assay
 Select ™ Salmonella Test
 Listeria species Test
 Listeria species Media
3M Microbiology

 3M Petrifilm Environmental Listeria Plate

Romer Labs

 AgraQuant®
Deoxynivalenol
(DON) Assay
Survey of
Performance Tested MethodsSM
 Approved methods come and go.
 Visit the AOACRI website to find out if a kit is
PTM approved
 http://www.aoac.org/testkits/testedmethods.ht
ml
Survey of
Official Methods of AnlaysisSM

Rapid microbiobiology methods

granted Official Methods of
Analysis since 2000
BioControl Systems, Inc
2002.07
SimPlate® Total Plate

2002.11
SimPlate® Yeast and
Mold Color Indicator
(Y&M-CI)

2005.03
Simplate® Coliform and E.
coli
BioControl Systems, Inc
2005.04
Assurance GDSTM for E. coli O157:H7
• 2005.05
Assurance GDSTM for Shigatoxin Genes
bioMérieux, Inc
 2001.07 VIDAS Immuno-Concentration Salmonella
Assay (ICS)
 2001.08 VIDAS Immuno-Concentration Salmonella
Assay (ICS) in Combination with Salmonella
Selective Plating Media, XLD
 2001.09 Salmonella in Foods, VIDAS
Immuno-Concentration Salmonella (ICS) /VIDAS
Salmonella Method, SLM
bioMérieux, Inc
 2001.09 Salmonella in Foods, VIDAS
Immuno-Concentration Salmonella (ICS) /VIDAS
Salmonella Method, SLM
 2004.02 VIDAS Listeria monocytogenes II (LMO2)
Immunoassay Method
 2004.03 VIDAS (SLM) Immunoassay
 2004.06 VIDAS® Listeria (LIS) Immunoassay
Dupont Qualicon
 2003.09
 BAX® System for the
Detection of
Salmonella

 2003.12
BAX® Automated
System for the
Detection of Listeria
monocytogenes
Neogen Corporation
• 2000.13
• REVEAL for Escherichia coli O157:H7
Screening Test

• 2007.02
• GeneQuenceTM Salmonella
Tecra
(Acquired by 3M)

 2000.07
TECRA Unique Salmonella Test

 2002.09
TECRA Listeria Visual Immunoassay
(modification)
3M Microbiology
• Petrifilm™

2000.15
Rapid Enumeration of
Coliforms

2001.05
Rapid S. aureus Count Plate
3M Microbiology
• Petrifilm™

2003.01
Enterobacteriaceae Count
Plate

2003.07, 2003.08 & 2003.11

Staph Express Count Plate
AOAC Validation
Completed

Thank You

www.aoac.org