Histoplasma capsulatum/Blastomyces dermatitidis, Molecular Detection, PCR
Method Description Describes how the test is performed and provides a method-specific reference
Following specimen processing, nucleic acids are extracted using the MagNA Pure Compact (Roche Applied Sciences). The extract is then transferred to a cuvette for amplification using the LightCycler real-time PCR platform (Roche Applied Sciences). The LightCycler is an automated instrument that amplifies and monitors the development of target nucleic acid (amplicon) after each cycle of PCR. The detection of amplicon is based of fluorescence resonance energy transfer (FRET), which utilizes hybridization probes. The presence of the specific organism nucleic acid is confirmed by performing a melting curve analysis of the amplicon. Primers and FRET hybridization probes were designed to target a 174-base pair (bp) region of the histidine kinase (DRK-1) gene of Blastomyces dermatitidis and a 192-bp region of the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene of Histoplasma capsulatum, respectively. The acceptor probe for Blastomyces dermatitidis was labeled with a Red-705 dye, while the acceptor probe for Histoplasma capsulatum was labeled with a Red-640 dye. Labeling the acceptor probes with 2 different dyes allows for simultaneous detection and differentiation of Blastomyces dermatitidis and Histoplasma capsulatum within a single PCR assay.(Unpublished Mayo method)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 Friday, 3 times per week
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.
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