A NEW TISSUE TECHNIQUE FOR EVALUATING EFFECTS OF BACILLUS-THURINGIENSIS TOXINS ON INSECT MIDGUT EPITHELIUM

Epithelial tissue wholemounts were produced after enzymatic removal of basal lamina and connective tissue from midguts of Trichoplusia ni la rvae. Wholemounts were nourished in artificial hemolymph and tissue vi ability was assessed for up to 24 hr using the vital dyes try-pan blue , acridine orange (AO), propidium iodide (PI), and 4',6-diamidino-2-ph enylindole (I)API), Peritrophic membrane synthesis and modification of Bacillus thuringiensis Cry1Ac protoxin to active toxin confirmed some normal epithelial function, Vital staining using the combination of A O and PI, or DAPI revealed altered membrane permeability in columnar e pithelial and regenerative cells of tissues treated with activated Cry 1Ac toxin while feeding and oral inoculation bioassays verified Cry1Ac toxicity. DAPI was selected to identify target cells in a rapid and h ighly sensitive assay. (C) 1997 Academic Press.