West Nile Virus, Molecular Detection, PCR
Method Description Describes how the test is performed and provides a method-specific reference
This LightCycler PCR assay has been optimized to detect common conserved sequences in the nonstructural protein of West Nile virus (WNV). Viral nucleic acid is extracted by the MagNA Pure automated instrument (Roche Applied Science) from cerebrospinal, or plasma. Primers directed to the nonstructural protein amplifies a specific sequence of the virus. For the test, WNV genomic RNA is transcribed to cDNA. The LightCycler instrument amplifies and monitors the development of target nucleic acid sequences after the annealing step during PCR cycling by fluorescence assay. This automated PCR system utilizes stringent air-controlled temperature cycling and capillary cuvettes to rapidly detect (30-40 minutes) amplicon development. 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. Analysis of the PCR amplification and probe melting curves are accomplished through the use of LightCycler software.(Cockerill FR III, Uhl JR: Applications and challenges of real-time PCR for the clinical microbiology laboratory. In Rapid Cycle Real-Time PCR Methods and Applications. Edited by U Reischel, C Wittwer, F Cockerill. Berlin, Germany, Springer-Verlag; 2002, pp 3-30)
PDF Report Indicates whether the report includes an additional document with charts, images or other enriched information
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; Continuously 7 a.m.-8 p.m. (June through November)
Monday, Wednesday, Friday; 6 a.m. (December through May)
Analytic Time Defines the amount of time it takes the laboratory to setup and perform the test. This is defined in number of days. The shortest interval of time expressed is "same day/1 day," which means the results may be available the same day that the sample is received in the testing laboratory. One day means results are available 1 day after the sample is received in the laboratory.
Monday through Thursday: 2 days Friday, Saturday: 3 days
Maximum Laboratory Time Defines the maximum time from specimen receipt at Mayo Medical Laboratories until the release of the test result
Specimen Retention Time Outlines the length of time after testing that a specimen is kept in the laboratory before it is discarded
Performing Laboratory Location The location of the laboratory that performs the test