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Unit Code 88909:
JC Virus DNA by PCR, Spinal Fluid

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Method Description

Viral nucleic acid is extracted from the specimen using the MagNA

Pure automated instrument (Roche Applied Science). Primers are

directed to the VP2 gene, which is a conserved sequence specific

for JCV. The LightCycler instrument (Roche Applied Science)

amplifies and monitors the development of target nucleic acid

sequences after the annealing step during PCR cycling. This

automated PCR system can rapidly detect amplicon development

through stringent air-controlled temperature cycling in capillary

cuvettes. The detection of amplified products is based on the

fluorescence resonance energy transfer (FRET) principle. For FRET

product detection, a hybridization probe with a donor fluorophore,

fluorescein, on the 3'-end is excited by an external light source and

emits light that is absorbed by a second hybridization probe with an

acceptor fluorophore, LC-Red 640, at the 5'-end. The acceptor

fluorophore then emits a light of a different wavelength that can be

measured with a signal that is proportional to the amount of specific

PCR product. (Whiley D, Mackay IM, Sloots TP:  Detection and

differentiation of human polyomaviruses JC and BK by LightCycler

PCR. J Clin Microbiol 2001;39:4357-4361)

Performing Laboratory Location

Rochester

Key