Detection of Low Levels of TNT Using Immunologic and Bioluminescent Techniques
Abstract
Several approaches toward the development of a bioluminescent immunoassay for TNT have been examined. Three basic types of immunoassays have been studied: (1) Homogeneous Bioluminescent Assay, (2) Heterogeneous Bioluminescent Assay, and (3) Amplified Heterogeneous Bioluminescent Assay. The homogeneous immunoassay involves preparing a TNP-luciferase conjugate which upon binding to anti-TNP antibody exhibits either a significant increase or decrease in luciferase activity, i.e., light production. A large change in light production was not observed and this approach was not pursued further. The heterogeneous immunoassay utilizes an antibody to TNP which has been covalently bound to a solid phase, i.e., Sepharose. The free TNT and TNP-luciferase compete for binding sites on the immobilized antibody and the bound luciferase-TNP is measured by light production. There is an inverse relationship between the amount of free TNT in the sample and the amount of TNP-luciferase bound. Using this procedure it was possible to detect 50 femtomoles of free TNT. The amplified heterogeneous assay is in principle the same as that described for the heterogeneous assay except the TNP is linked to glucose-6-phosphate dehydrogenase rather than luciferase. This enzyme has a large turnover number and therefore one molecule of glucose-6-phosphate dehydrogenase can produce 70, 000 molecules of product per minute.
Document Details
- Document Type
- Technical Report
- Publication Date
- Dec 01, 1982
- Accession Number
- ADA130090
Entities
People
- Daniel Vellom
- Marlene Deluca
Organizations
- University of California, San Diego