Dot blot enzyme-linked immunosorbent assay for monitoring the fate of insecticidal toxins from Bt in soil
Authors: Tapp, H., Stotzky, G.
Journal, Volume, Year: Applied and Environmental Microbiology, volume 61(2):602-609, 1995
Toxins from Bt kurstaki and tenebrionis were isolated and antibodies against each toxin were prepared. An assay using dot-blot procedures was developed. Samples were prepared by adding the toxins to clay, soil, or clay-amended soil. Soils were incubated for different amounts of time, followed by fractionation of the soil into clay, silt, and sand. Qualitative determination of toxin was accomplished by spotting soil or clay onto a protein-binding PVDF membrane. The membrane was developed by ELISA, ending in a darkened spot on the membrane if the toxin was present. Lower detection limits were approximately 3 ng regardless of the matrix (bound to soil, clay, or free). Quantitative data was not possible due to dislodging of particles from the membrane during ELISA. Toxins were more likely to be detected in the clay fraction of soils as compared to sand or silt. Incubation of soils in nonsterile conditions indicated that toxins bound to clay degraded slowly and were still detectable after 40 days. Insect bioassays were more sensitive than the dot-blot assay.
This method is fast, fairly sensitive, and low cost. However, the insect bioassays are more sensitive. This test is only qualitative or at best semi-quantitative.