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" Development of porcine-specific monoclonal antibodies and their application in the detection of pork in cooked meat products "
F.-C. Chen
Y. H. P. Hsieh
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Latin Dissertation
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| Language of Document
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English
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| Record Number
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1112962
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| Doc. No
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TLpq304427687
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| Main Entry
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F.-C. Chen
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Y. H. P. Hsieh
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| Title & Author
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Development of porcine-specific monoclonal antibodies and their application in the detection of pork in cooked meat products\ F.-C. ChenY. H. P. Hsieh
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| College
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Auburn University
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| Date
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1998
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| student score
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1998
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| Degree
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Ph.D.
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| Page No
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133
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| Abstract
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Methods for detection of meat adulteration are required for implementation of food labeling regulations and product quality control. The use of monoclonal antibody (MAb) in immunoassay would provide a standardized and cost-effective method for identifying undeclared species in meat products. The purpose of this research was to develop MAb-based enzyme-linked immunosorbent assays (ELISAs) for the detection of pork in cooked meat products. In the first study, four hybridoma cell lines (5H9, 5H8, 2E2, and 8A4) secreting MAbs specific to porcine thermal-stable muscle protein (TSMP) were produced. All of these MAbs showed a strong reactivity to pork extract without noticeable cross-reactivity to meat extract from beef, lamb, horse, deer, chicken, turkey and duck as determined by indirect ELISA. Western blotting revealed that these MAbs reacted with three protein bands (20.5, 22 and 24 kD) from raw pork extract: the protein band (24 kD) that was also presented in cooked pork extract was identified as porcine-specific TSMP. Epitope analysis indicated that the four MAbs recognized same or closely located antigenic sites on the pork TSMP. Indirect ELISA using 5H9 ascites fluid enabled the detection of 1% pork (w/w) in raw and cooked heterogenous meat mixtures. The second study was conducted to improve analytical validity of indirect ELISA for detection of pork in cooked meat products and to evaluate quantitative aspect of the assay for estimating pork content in meat samples. The ELISA developed using purified 5H9 immunoglobulin G was able to detect as low as 0.5% (w/w) of pork in heterogenous meat mixtures. The assay appeared to be skeletal-muscle-specific with no cross-reactivity to proteins from pork cardiac muscle, smooth muscle, serum and other organs. Overall intra-assay and inter-assay coefficients of variation were 5.5% and 7.9%, respectively. Accuracy of analyzing market samples was 100% as verified by the product labeling and confirmed by a commercial meat speciation test. Quantitative determination of adulteration levels was not different (P > 0.05) for pork from various retail cuts. However, non-meat protein additives, sample matrix, pH, salt concentration, and cooking temperature appeared to be factors influencing accurate prediction of pork content. Results suggest that a matching matrix is required for standard calibration in the quantitative estimation. The developed ELISA proved to be a reliable assay for sensitive detection of pork in various cooked meat products. The success in producing porcine-species MAbs and identifying TSMP implies a promising future in developing specific MAbs for detection of other meat species.
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Biological sciences
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Food science
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meat adulteration
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porcine
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