A sensitive bioluminescent immunoassay for dinitrophenol and trinitrotoluene

✍ Scribed by Jon Wannlund; Marlene Deluca

• Publisher: Elsevier Science
• Year: 1982
• Tongue: English
• Weight: 771 KB
• Volume: 122
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(82)90300-1

✦ Synopsis

A bioluminescent immunoassay for measuring dinitropbenol and trinitrotoluene (TNT) has been developed. The DNP and TNT were covalently linked to firefly luciferase. resulting in a conjugate containing 1 mol of DNP or trinitrophenyl (TNP) per mole of luciferase. The conjugate retained 90% of its original catalytic activity. When the conjugate was incubated with immobilized anti-TNT or anti-DNP and varying concentrations of free TNT or DNPleucine, the amount of conjugate bound was inversely proportional to the concentration of the free compound. Using this procedure it is possible to detect 2.5 pmol of DNP-leucine and 1 .O pmol of TNT. If the TNP or DNP is linked to glucose-6-phosphate dehydrogenase instead of luciferase, much lower quantities of antigen can be detected. This is due to the fact that this enzyme has a large turnover number so that amplification is possible. The NADH produced is measured using immobilized bacterial NADH:FMN oxidoreductase and luciferase. With this procedure, 10 amol (10-l' mol) of antigen can be measured. These procedures should be suitable for measuring any antigen.