Cytomegalovirus, Molecular Detection, Quantitative PCR, Plasma
Method Description Describes how the test is performed and provides a method-specific reference
For this assay, viral DNA is extracted from 0.2 mL of plasma by the MagNA Pure automated instrument (Roche Applied Science). LightCycler PCR primers and probes have been designed to detect target cytomegalovirus (CMV) DNA in the UL54 gene of the virus. The LightCycler instrument amplifies and monitors target nucleic acid sequences by fluorescence during PCR cycling. This is an automated PCR system that can rapidly detect amplified product development. The detection of amplified products is based on the 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, which 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 light of a different wavelength that is measured with a signal that is proportional to the amount of specific PCR product. Quantitative results are obtained by incorporating known copy levels (5 standards) of CMV DNA (plasmid) into the assay, together with the specimen. LightCycler instrument software calculates the level of CMV DNA (copies/microliter) in the specimen from a standard curve of the quantitative levels of the reference plasmid CMV DNA. The process is completed in a closed tube system. (Unpublished Mayo method)
Day(s) and Time(s) Test Performed Outlines the days and times the test is performed. This field reflects the day and time the sample must be in the testing laboratory to begin the testing process and includes any specimen preparation and processing time required before the test is performed. Some tests are listed as continuously performed, which means assays are performed several times during the day.
Monday through Saturday; Varies