GeneQuence utilizes a novel DNA hybridization technology which assays for Salmonella, Listeria spp., Listeria monocytogenes, and Campylobacter. Each test kit uses two specific DNA elements ensuring the highest of specificity, thereby increasing your confidence of the rapid results. Each kit has been developed to detect 1-5 CFU/25g sample in less than 2 hours. By coupling this easy to use technology with an automated plate handling unit, it is possible to test more than 450 samples in an 8 hour work day with very little hands on time. Contact us if you are ready to simplify your pathogen testing program.
First, a portion of the enrichment culture is placed into a test tube. A lysis reagent is added, which disrupts the cell and releases the nucleic acid target molecules. A portion of the lysed sample is then transferred to a microwell and the probe reagents are added. (Figure 1.)
The probe reagents consist of: 1) an oligonucleotide capture probe specific to rRNA sequences of the target organism and labeled at the 3’ end with polydeoxyadenylic acid (poly dA); and 2) an oligonucleotide detector probe also specific to rRNA sequences of the target organism and labeled at the 5’ end with the enzyme horseradish peroxidase (HRP). The hybridization reaction is then allowed to proceed for one hour. If target rRNA is present in the sample, both probes will hybridize to their complementary sequences on the target molecule.
The resulting complex is captured onto the solid phase coated with polydeoxythymidylic acid (poly dT), which is complementary to the poly dA portion of the capture probe. Unbound probe is then washed away, and a substrate of HRP is added.
Following a short incubation, blue color indicates the presence of hybridized detector probe in the complex and thus the presence of rRNA from the target organism. Stop solution is added to convert blue to yellow color and results are determined spectrophotometrically at 450 nm. An absorbance value in excess of a predetermined threshold indicates a positive test result. (Figure 2.)
GeneQuence Method versus PCR-based Tests
|Point of Comparison||GeneQuence||PCR||Benefit of GeneQuence|
|Assay time||1 hr, 50 min||3 hrs, 30 min (average)||Faster results. GeneQuence also has at least 25% less off-line labor time (instrument set-up, manual reagent addition, etc.) than PCR methods.|
|Commodities||Good performance across foods||Some food types difficult||Published data documents problems PCR in certain food commodities.|
|Flexibility||2-4 pathogens at same time||1 pathogen per run||GeneQuence, due to its superior automation, allows 2-4 pathogens to be assayed concurrently. To do this by PCR, more than one instrument would be needed (at a higher capital cost).|
|Processing||2-4 96-well plates concurrently||1 set of 96 samples at a time||As soon as one plate is finished, GeneQuence can accept another sample set. PCR systems must wait until the entire run is complete before loading another set. GeneQuence can provide uninterrupted, continuous operation.|
|Technology||Nucleic acid hybridization||PCR||The thermostable polymerase used in PCR has documented inhibition problems that lead to false negatives with several matrices. GeneQuence avoids this issue entirely by using nucleic acid hybridization. GeneQuence also probes the target directly, rather than an intermediate amplified from the original target.|
|Throughput||1.32 samples per minute||0.46 samples per minute||187% more throughput on GeneQuence will minimize retention time of inventory.|
|Point of Comparison||GeneQuence||ELISA||Benefit of GeneQuence|
|Detection||rRNA||Protein||1,000 to 10,000 copies of rRNA per cell give GeneQuence its high level of sensitivity.|
|Inclusivity||DNA Hybridization||Sandwich ELISA||Allows detection of all serovars of Salmonella|
|Pathogen Target||rRNA||Protein||RNA is very short-lived as it is constantly broken down by ubiquitous RNases. Couple that with dilutions from enrichment, GeneQuence will not detect dead bacteria.|
|Procedure||Manual or Automated||Varies||Some ELISA systems can only be run on an instrument. Others specify only a manual method that may be adapted by the user into an automated system. Only GeneQuence allows the user to perform the test manually or in a company-supported automated format.|
|Two levels of stringency (binding of target rRNA to capture probe, and rRNA to detector probe) decrease false positive rates with GeneQuence.|
|Throughput||DSX||Other platforms||The DSX can process more than 450 samples in an 8-hour shift, at least double that of any ELISA system on the market. This will reduce labor and equipment costs.|