Detection and bioactivity of Cry1Ab fragments: Implications on risk assessments

Use of transgenic crops expressing genes for the production of insecticidal crystalline (Cry) proteins continues to increase; however, concerns that these proteins may have adverse effects on non-target organisms persist. Accurate detection of these proteins in various environmental matrices is essential for determining possible exposure of non-target organisms to these proteins. Enzyme-linked immunosorbent assays (ELISAs) have been used widely for detection of Cry proteins in the environment.  A major obstacle to this approach is that their results are not typically validated biologically to ensure that only bioactive Cry proteins are detected. Therefore, concentrations of the bioactive protein may not be accurately represented in exposure assessments. This could potentially lead to overly conservative risk assessments and unnecessary regulation. Thus, in order to properly study Cry proteins in environmental matrices there is a need for standardized detection methods that are biologically validated. This research project will improve methods of detection for Cry proteins in environmental matrices by developing a framework for biological validation of ELISA procedures. Proteinase K was used to generate a fragment of Cry1Ab that is around 5 kDa in size. The fragments will then be analyzed with ELISAs to determine if fragments are detectable, and with bioassays to determine if the fragments retain bioactivity. Finally, specific guidelines for biological validation of ELISAs will be suggested.