DEVELOPMENT OF A FLOW-INJECTION ANALYSIS (FIA) IMMUNOSENSOR FOR THE DETECTION OF ESCHERICHIA-COLI

A flow injection immunoanalysis (FIA) system has been developed for th e detection of Escherichia coli in artificially contaminated food samp les. Anti-E. coli antibodies were covalently immobilized onto porous a minopropyl glass beads via glutaraldehyde activation to form an immuno reactor. After adsorption of the cells onto anti-E. coli antibody boun d glass beads, 4-methylumbelliferyl-beta-D-glucuronide was injected in to the system which was then hydrolyzed by the adsorbed E, coli cells containing beta-D-glucuronidase, an enzyme which is very specific to E . coli and to a few other strains of Shigella. Fluorescent 4-methylumb elliferone released from the enzymatic reaction was then detected by a fluorometer. Owing to the specificity of the antibody towards E. coli , the FIA system was very selective for detection of E. coli whereas S higella boydii, another GUD-positive bacterium, did not give any respo nse. The FIA system was successfully used for detecting as low as 5x10 (7) CFU/ml E, coli in less than 30 min and was reusable for at least 3 00 repeated assays. The immunoreactor yielded reproducible results dur ing 3 months of experimentation if stored overnight at 4 degrees C in carrier buffer containing 0.05 to 0.25% Tween 20.