Microtiter-based assay for evaluating the biological activity of ribosome-inactivating proteins.
A microtiter assay was developed to quickly measure the biological activity of ricin or other ribosome-inactivating proteins. Nuclease-treated rabbit reticulocyte lysate containing luciferase mRNA was used to measure toxin activity via inhibition of protein synthesis. The relative biological activity was determined by comparing luminescence levels in treated samples versus those of untreated controls. The amount of luciferase translated, as measured by luminescence, was inversely proportional to the toxin concentration. Linear dose response curves were generated for both class I and class II toxins. When compared to normal serum controls, specific antibody against ricin effectively inhibited ricin activity. The assay is suitable for efficient in vitro screening of therapeutics, as well as for identifying samples containing ribosome-inactivating proteins.
Hale
M L
ML
Toxinology Division, United States Army Medical Research Institute of Infectious Diseases, Fort Detrick, MD 21702 5011, USA. martha.hale@det.amedd.army.mil
D000818:Animals; D001681:Biological Assay / Q000379:methods; D002474:Cell-Free System; D004307:Dose-Response Relationship, Radiation; D006358:Hot Temperature; D066298:In Vitro Techniques; D008156:Luciferases / Q000235:genetics; D008163:Luminescent Measurements; D010940:Plant Proteins / Q000187:drug effects* / Q000378:metabolism; D014176:Protein Biosynthesis; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011817:Rabbits; D012156:Reticulocytes / Q000378:metabolism; D012260:Ribonucleases / Q000378:metabolism; D012269:Ribosomal Proteins / Q000037:antagonists & inhibitors*; D012270:Ribosomes / Q000378:metabolism*; D012276:Ricin / Q000494:pharmacology*; D012680:Sensitivity and Specificity; D013997:Time Factors; D014118:Toxins, Biological / Q000145:classification / Q000494:pharmacology / Q000633:toxicity
D010940:Plant Proteins; D011500:Protein Synthesis Inhibitors; D012333:RNA, Messenger; D012269:Ribosomal Proteins; D014118:Toxins, Biological; D012276:Ricin; D008156:Luciferases; D012260:Ribonucleases