The document discusses quality control for molecular diagnostics including maintaining nucleic acid integrity during sample collection and transport, establishing contamination control through unidirectional workflow, and using positive and negative controls. It also describes real-time PCR processes, showing amplification curves for positive and negative samples, and examples for influenza A subtyping. A risk of in-house PCR is mentioned as using non-optimized primers and controls requiring cautious result interpretation.

1. 1
Quality Control:
Molecular Diagnostics

2. Molecular Diagnostics Quality Control - Module 8 2
2
Overview
 Sample Collection, Transport, and Processing
 maintain Nucleic acid integrity through these
processes
 Contamination Control
 establish a unidirectional work flow from a
DNA-free area for reagent preparation, to a
sample processing area, to area where
amplification and detection occur
 Positive and Negative Controls
 Reference: CLSI MM3-A (Molecular Diagnostic
Methods for Infectious Diseases)

3. Molecular Diagnostics Quality Control - Module 8 3
3
Quality Control:
Molecular Diagnostics

4. Molecular Diagnostics Quality Control - Module 8 5
5
Real Time PCR
SAMPLE
PRIMER
Taq polymerase
Amplified
Target NA
SYBR Green
Quantitative
Fluorescence
+ - In

5. Molecular Diagnostics Quality Control - Module 8 6
6
Amplification Reactions
Amplification / Time
POSITIVE
NO AMPLIFICATION

6. Molecular Diagnostics Quality Control - Module 8 7
7
Real Time RT-PCR

7. Molecular Diagnostics Quality Control - Module 8 8
8
A/H3

8. Molecular Diagnostics Quality Control - Module 8 9
9
A/H5

9. Molecular Diagnostics Quality Control - Module 8 11
11
Risks with PCR
 most RT-PCR is still done as “in-
house” or “home-brew”
primers are not necessarily optimized to
test
positive controls are “in-house”
negative controls are “in-house”
 interpretation of tests need to be
done with caution