Novel Ultrasensitive Colorimetric Assay for DNA

Award Information
Agency: Department of Health and Human Services
Branch: N/A
Contract: N/A
Agency Tracking Number: 22089
Amount: $749,534.00
Phase: Phase II
Program: SBIR
Awards Year: 1995
Solicitation Year: N/A
Solicitation Topic Code: N/A
Solicitation Number: N/A
Small Business Information
Elcatech, Inc.
64-1001 S. Marshall Street, Winston-salem, NC, 27101
DUNS: N/A
HUBZone Owned: N
Woman Owned: N
Socially and Economically Disadvantaged: N
Principal Investigator
 Mark X. Triscott
 (919) 777-3624
Business Contact
Phone: () -
Research Institution
N/A
Abstract
The detection of specific DNA sequences for the diagnosis of genetic diseases, cancer, and infectious disease is becoming increasingly widespread. A number of innovative techniques have been employed, including the oligonucleotide ligation assay and reverse dot-blot hybridization. The end-point, and to a certain extent the effectiveness of these assays, is determined by the sensitivity of the DNA detection techniques used. Progress has been significantly enhanced through the use of the polymerase chain reaction (PCR). However, implementation of PCR can be expensive as well as equipment and expertise intensive. We have applied a technique with proven effectiveness in the field of immunoassays for the non-isotopic detection of DNA; initial data indicates an increase in sensitivity 10(2)-10(3) times higher than conventional techniques. The assay, which we have called EDNA-ELCA, uses the amplification of the coagulation cascade to produce a calorimetrically detectable end product. A DNA template is labelled with biotin and a haptene, and is then immobilised in a streptavidin coated microtiter plate. An antibody against the haptene is conjugated to a snake venom coagulation activator (RVVXa), and introduced into the microtiter well. The assay is then developed by the addition of coagulation factors, including an enzyme labelled fibrinogen. The deposition of enzyme labelled fibrin is indicative of a positive reaction. We have chosen three specific applications to prove the utility of this method. They are detection of the deltaF508 mutation of cystic fibrosis, a codon 12 mutation in the first exon of K- ras for colon cancer, and the mecA gene in methicillin resistant Staphlococcus aureus.

* information listed above is at the time of submission.

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