Enzyme-linked Immunoassay for the Detection and Quantification of Cry Protein in Bacillus thuringiensis Strains

Bacillus thuringiensis (Bt), which produces cry protein during its sporulation phase, is a commonly used biopesticide in the market to control a wide range of insect pests belonging to Coleoptera, Lepidoptera, and Diptera. The development of assays for detecting and quantifying cry protein to commercialize biopesticides is in high demand. Various methods are available to detect the cry protein; however, enzyme-linked immunosorbent assays (ELISA) are the preferred method to quantify the cry protein as it is highly sensitive, reliable, specific, cost-effective, easy to use, and give rapid results. Our study aimed to develop indirect ELISA to quantify cry proteins of in-house Bt isolates and the standard strains of Bt var kurstaki. In this study, the method for isolating the cry proteins was optimized. The isolated protein concentration was estimated using Bradford's method. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis showed the banding pattern 140 KDa and 65 KDa for cry1ac and cry2ab proteins, respectively. The Dot blot assay showed the presence of cry protein in all the in-house Bt isolates. The newly developed indirect ELISA quantified the cry proteins using the curve expert standard..