30C-13 |
Monoclonal antibodies against troponin I for the detection of mammalian meats in poultry products |
F. C. CHEN, Department of Nutrition and Food Science, Auburn University, 328 Spidle Hall, Auburn, AL 36849 and Y. H. P. Hsieh.
Immunoassays for species authentication of meats are essential tools for quality assurance and labeling regulations. We have demonstrated the feasibility of troponin I (TnI) as a thermal-stable antigen for species identification of heat-processed meats. The objectives of this study were to produce and characterize monoclonal antibodies (Mabs) specific to TnI, and to develop a Mab-based enzyme-linked immunosorbent assay (ELISA) for detection of mammalian meats in cooked poultry products. Purified TnI from bovine skeletal muscle was used as immunogen for Mab production. Specificity of Mab was characterized by immunoblot with TnI from different species. Epitopes of Mabs were mapped by a competition assay between a pair of biotin-labeled and unlabeled Mabs. An ELISA employing these Mabs was developed and tested on artificially adulterated chicken and turkey samples. Three IgG1-subclass Mabs (1F9, 2G3, 8A12) with unique specificity to TnI were selected from more than 4000 hybridoma clones. Mab 1F9 reacted to both cardiac and skeletal TnI of mammalian and poultry species, while Mab 2G3 recognized only skeletal TnI. With Mab 8A12, it was possible to differentiate skeletal TnI of mammalian origin (pig, cattle, sheep, horse, deer) from poultry (chicken, turkey, duck, goose). Epitope comparison revealed that little competition existed between two of the different Mabs (less than 10 % inhibition) at concentrations of 1 to 8 fold excess of one another. Under similar conditions, a pair of the same Mab yielded 47 to 88% inhibition. Results suggested that these Mabs bind to non-overlapping antigenic sites on the TnI molecule. A sandwich ELISA utilizing Mab 8A12 and 2G3 as paired capture reagents was developed. The assay was capable of determining levels as low as 0.5% of mammalian meats in cooked ground chicken or turkey. These TnI specific Mabs would be excellent probes in studying muscle chemistry and physiology. The sandwich ELISA demonstrated usefulness of these Mabs in a rapid and cost-effective screening test for the detection of species adulteration in cooked poultry products.
Session 30C, Muscle Foods: Tenderness, Quality, Processing, Marination, Oxidation, and Shelf-Life
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