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Laboratory Testing for Hepatitis C



Hepatitis C: Molecular Assays

Slide 11

April 2008

I want to spend a few minutes describing the molecular methods that are currently used commercially for qualitative and quantitative molecular tests.
 
In the first column in the left, you see a polymerase chain reaction (PCR), which amplifies the viral nucleic acid target. In this case, it’s HCV RNA but because PCR is a DNA-target amplification process and the virus is an RNA virus, it is necessary to have the first step of reverse transcription by making a complimentary DNA copy of the viral RNA target. For HCV RNA amplification by PCR, it’s really a reverse transcription PCR (RT-PCR). From the complimentary DNA copy, one can generate multiple copies of subsequent DNA.

With transcription mediated amplification (TMA) for HCV RNA the viral target is copied by reverse transcription to make complimentary DNA, and then using transcription the complimentary DNA is used to make multiple copies of RNA. Both of these methods are amplifying the viral genome, so it’s a target amplification method.

The last method which is used commercially is the branch DNA (bDNA) method. Here, the viral RNA target is not amplified, but it is captured by oligonucleotide probes that are specific for the region of the HCV RNA gene. The probe contains multiple attachment sites for signal reagents to be attached, so a single viral target could be captured and the signal is amplified to indicate the presence or absence of the target. So here, you see this is a signal amplification method as opposed to a target amplification method.

Molecular Assays

 


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