Virology and Molecular Diagnostics
This domain tests specimen transport for viruses and the contamination controls that make a molecular result trustworthy.
Specimen handling
- Use viral transport medium with antibiotics and flocked nylon swabs; avoid calcium alginate and dry swabs. Transport at 2–8 °C and freeze at −70 °C if delayed; avoid repeated freeze-thaw.
- Viral load testing requires EDTA plasma (HIV-1, CMV); whole blood for CMV DNA; CSF for herpes simplex PCR; serum for serology.
- Culture is slow and now largely replaced by NAAT for routine diagnosis.
Molecular workflow
- Unidirectional workflow: reagent preparation, extraction, and amplification in separate rooms with dedicated equipment and moving away from clean areas.
- Every run needs a negative control, a positive control, and an internal/extraction control to detect inhibition.
- Prevent carryover with aliquoted reagents, uracil-N-glycosylase with dUTP master mixes, and bleach cleanup.
- Ct values are inversely proportional to target amount; report only within the validated range and limit of detection.
Worked example
A run's negative control amplifies at Ct 36 while a patient sample amplifies at Ct 34. Reasoning: any signal in the negative control indicates contamination, so the run is invalid and the patient result cannot be reported even though the Ct is plausible. Decontaminate surfaces, open fresh reagents, and repeat the batch with new controls and the patient sample.
Traps
Reporting a low-positive result without repeat testing; using serum instead of EDTA plasma for viral load; sending a swab in bacterial transport medium; and treating a Ct value as quantitative when the assay is only qualitative.