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The Gen-Probe AccuProbe system uses an acridinium-labeled, single-stranded DNA probe that is complementary to the ribosomal RNA of the target organism. After the ribosomal RNA is released from the organism, the labeled DNA probe combines with the target organism's ribosomal RNA to form a stable DNA:RNA hybrid. Chemiluminescence is used as an indicator to detect specific hybrids.(Musial CE, Tice LS, Stockman L, Roberts GE: Identification of Mycobacterium from culture by using Gen-Probe rapid diagnosis system for Mycobacterium avium complex and Mycobacterium tuberculosis complex. J Clin Microbiol 1988;26:2120-2123)
The DNA sequence analysis utilizes a 500 basepair region of the 16S rRNA gene as the target for identification of mycobacteria and is performed using the MicroSeq kit from Applied Biosystems. Sequence data generated is compared to several different databases of known mycobacterial and aerobic actinomycete sequences to obtain organism identification. These include MicroSeq, NCBI GenBank, and Mayo Clinic Mycobacteria database. A 100% or greater agreement with a database strain is needed for an acceptable identification to the species level.(Hall L, Doerr KA, Wohlfiel SL, Roberts GD: Evaluation of the MicroSeq system for identification of mycobacteria by 16S ribosomal DNA sequencing and its integration into a routine clinical mycobacteriology laboratory. J Clin Microbiol 2003;41:1447-1453)
MALDI-TOF mass spectrometric analysis is done using the Bruker BioTyper platform and the Bruker BDAL library, Bruker Mycobacterial Library and the Mayo Clinic Library. A spectral score of > or =2.0 is required for identification to the species level.
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