Standardization of NAT Extraction Methods

1
Standardization of NATExtraction Methods

• Paul D. Neuwald, PhD
• AcroMetrix Corporation, Benicia, CA
• SoGAT XVIII
• Bethesda, MD, USA
• May 24, 2005

2
Standardization of NAT Extraction MethodsOverview

• Potential sources of variation in NAT assays
• Specimen type
• Platform / Procedure
• Operator / Laboratory
• Calibrators / Standards
• Assay reagents
• Extraction method

3
Standardization of NAT Extraction MethodsOverview

• Potential sources of variation in NAT assays
• Specimen type
• Platform / Procedure
• Operator / Laboratory
• Calibrators / Standards
• Assay reagents
• Extraction method

4
Example CMV Assay Comparison StudyStudy Design

• Each participating laboratory
• Extracted CMV positive reference panel members
  using its standard method for clinical specimens
• Tested the panel members as unknowns in
  replicates of 4 using its standard CMV DNA assay
  method and calibrators
• Returned quantification results for analysis and
  comparison to results from other labs (N 23)

5
Example CMV Assay Comparison StudyReference
Panel

• OptiQuant CMV DNA Panel

Panel Member Nominal CMV DNA Copies/mL
0 0
1 500
2 5,000
3 50,000
4 500,000
Value assignment based upon quantification using
the COBAS AMPLICOR CMV MONITOR Test
6
CMV DNA Assay Comparison StudyInterlaboratory
Comparison

• Overall Results for 23 Laboratories

7
CMV DNA Assay Comparison StudyInterlaboratory
Comparison

• Examples from 3 Individual Laboratories

8
Methods used in CMV DNA Assay Study
9
CMV DNA Assay Comparison StudyInterlaboratory
Comparison

• LightCycler Different Extraction Methods

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CMV DNA Assay Comparison StudyInterlaboratory
Comparison

• ABI 7000 Series Different Extraction Methods

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Parvovirus B19 DNA AssayExtraction Comparison
12
Standardization of NAT Extraction Methods Future
Considerations

• Need for better controlled collaborative studies
  to compare extraction methods for
• Different specimen types / calibrators
• Different platforms / instruments / procedures
• Different assay methods / reagents
• Different operators / laboratories
• Initially, conduct a study that focuses on
  comparing extraction methods for one specimen
  type, on one platform, using common calibrators
  and reagents
• AcroMetrix is willing to coordinate such a study

13
Acknowledgments
AcroMetrix Corporation Association for Molecular
Pathology
Laboratories Participating in the CMV DNA Assay
Comparison Study
Advocate Lutheran General Hosp Childrens Mercy
Hosp Baylor University Med Center Carolinas
HealthCare System Childrens Hosp Loma Linda
Mercantile Mayo Clinic Medical College of
Virginia Medical University of South
Carolina Medicine University of Rochester Med
Center Molecular Pathology Network, Inc Oregon
Health Sciences University
Sharp Memorial Hosp Spectrum Health St Barnabas
Med Center The Methodist Hosp The Nebraska Med
Center UMDNJ/NJ Medical School University of
Florida University of North Carolina University
of Pitts/Magee Womens Hosp University of Texas
SW Med Center University of Washington Med
Center Washington Hosp Center