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The Great Basin Toxigenic C. difficile Direct Test, performed on the Great Basin PA500 Analyzer, is a qualitative in vitro diagnostic test for the detection of toxigenic Clostridioides difficile in unformed (liquid or soft) stool samples collected from patients suspected of having C. difficile infection (CDI). The automated assay utilizes polymerase chain reaction (PCR) to detect a conserved region of the toxin gene (tcdB) associated with toxin-producing C. difficile. The Toxigenic C. difficile Direct Test is intended for use as an aid in the diagnosis of CDI in conjunction with clinical and epidemiological risk factors.

The Great Basin Toxigenic C. difficile Direct Test (CDF2) performed on the PA500 Analyzer utilizes automated, hot-start PCR amplification technology to amplify specific nucleic acid sequences that are then detected using hybridization probes immobilized on a modified silicon chip surface, in a single-use, self-contained test cartridge. A swab volume of the specimen (raw stool) is first processed using the Sample Preparation Device (SPD). During processing through the SPD the specimen is infused with a synthetic bacterial sample processing control (SPC). An aliquot (250 µL) of the eluate obtained from the SPD, containing diluted specimen mixed with SPC, is loaded into the sample port of the CDF2 Test Cartridge. Genomic DNA is extracted from microbial cells and diluted to reduce potential inhibitors of PCR. During the PCR process, biotin-labeled primers direct the amplification of specific nucleic acid sequences within a conserved region of the C. difficile Toxin B (tcdB) gene. Following PCR, biotin-labeled, amplified target DNA sequences are hybridized to sequence specific probes immobilized on the silicon chip surface and incubated with antibody conjugated to the horseradish peroxidase enzyme (HRP). The unbound conjugate is washed away, and then tetramethylbenzidine (TMB) is added to produce a colored precipitate at the location of the probe/target sequence complex. The resulting signal is detected by the automated PA500 Optical Reader within the PA500 Analyzer System. The SPC undergoes the same extraction, and detection steps as the sample in order to monitor for inhibitory substances, as well as process inefficiency due to instrument or reagent failure. No operator intervention is required once the sample is loaded into the sample port and the Toxigenic C. difficile Direct Test cartridge is loaded into the PA500 Analyzer. The Great Basin System is a fully automated in vitro diagnostic system that includes: the PA500 Analyzer, single-use SPDs, single-use Toxigenic C. difficile Direct Test Cartridges, and the PA500 Data Analysis Software Program. The PA500 Analyzer is designed to perform automated sample preparation, target amplification via PCR, chip-based (optical) target detection, and integrated data analysis in less than two hours.