UNIQUE IMMUNOASSAYS FOR DETECTION OF CIGUATOXIN AND RELATED MARINE TOXINS USING MONOCLONAL ANTIBODIES

THIS PROPOSAL DESCRIBES A UNIQUE AND HIGHLY SIMPLIFIED IMMUNOLOGICAL PROCEDURE USING A MONOCLONAL ANTIBODY TO CIGUATOXIN IN A TWO-STEP PROCEDURE. THE MONOCLONAL ANTIBODY(MAB) IS PREPARED BY THE MOUSE HYBRIDOMA TECHNIQUE, USING HIGHLY PURIFIED CIGUATOXIN CONJUGATED TO PROTEIN CARRIERS. THE MAB-CTX THUS OBTAINED IS CONJUGATED TO COLLOIDAL GOLD (< 0.45 UM DIAMETER) OR COLORED LATEX PARTICLES. THIS WILL BE THE SPECIFIC DETECTION REAGENT. TOXINS IN FISH SAMPLES WILL BE DETECTED BY TOUCHING THE FISH TISSUE WITH BAMBOO STICK COATED WITH ORGANIC SOLUBLE LIQUID CORRECTION FLUID. THE STICK IS THEN FIXED FOR ONE SECOND IN ABSOLUTE METHYL ALCOHOL, AIR-DRIED, AND IMMERSED IN THE COLORED SUSPENSION OF MAB-CTX COUPLED TO GOLD OR LATEX SUSPENSION. A STICK THAT SHOWS COLOR WITHIN A FEW MINUTES WILL BE CONSIDERED POSITIVE. THE LIQUID PAPER CORRECTION FLUID COAT HAS BEEN PREVIOUSLY EXAMINED IN THE STICK ENZYME IMMUNOASSAY AND APPEARS TO ATTRACT CIGUATOXIN AND RELATED POLYETHERS FROM FISH TISSUES. THE FEATURES OF THIS "SOL PARTICLE IMMUNOASSAY" WILL MAKE IT USABLE UNDER VARIOUS CONDITIONS (LABORATORY, ON SHIP WHILE REEF FISHING, OR AT HOME). A CONVENTIONAL COMPETITIVE ENZYME IMMUNOASSAY FOR ASSAYING MARINE TOXINS (ANTIGEN) WILL ALSO BE ESTABLISHED.