15C-3 |
Properties and characterization of a monoclonal antibody for its use in screening Bacillus endospores in skim milk powder |
S. FULLER, Dairy Products Technology Center, California Polytechnic State University, 1 Grand Ave., Bldg 18-2, San Luis Obispo, CA 93407 and R. R. Jiménez-Flores. Finding a rapid detection method for endospores in skim milk powder (SMP) is important to the production of a safe and marketable product. Endospores are unique in their ability to resist the thermal processing of SMP and are able to germinate and grow initiating proteolysis and lipolysis. Bacillus cereus poses a potential health risk given that some strains are pathogenic. Current detection of endospores in milk powder is time consuming, inefficient and inaccurate. Therefore, a rapid, more efficient method is needed. Our objectives were to develop a rapid detection method using a monoclonal antibody (developed and donated from North Carolina State University). The monoclonal antibody binds to proteins on the exosporium of the endospore coat. Both ELISA and western blots were used to characterize the antibody interactions thoroughly. Through these immunochemical assays we were able to study the specificity of this antibody. We found the antibody to be highly selective to B. cereus and 5 other detrimental Bacillus strains found in SMP. Optimization of this method was necessary upon the discovery that commercial secondary antibodies used as reporters have, in some cases, strong cross reactivity. F(ab) based secondary antibodies were found to strongly react with a protein on the exosporium with a MW around 28kDa. The epitope is a large (>212kDa) polymer, likely a glycoprotein. Our optimized method can detect spore loads very accurately between 103 and 105, in some cases as low as 102. Results can be obtained in ~6-8 hours using these assays. This sensitive assay has proven to be superior to current practices in the detection/identification of endospores in SMP.
Session 15C, Dairy Foods
|