Differentiation of bovine from porcine gelatines using polyclonal anti-peptide antibodies in indirect and competitive indirect ELISA.
Gelatine is a collagen derivative obtained from bones and hides/skin mainly from bovine and pigs. As a consequence of the outbreak of bovine spongiform encephalopathy (BSE), the use of bovine gelatine in feed, food and pharmaceutical products has been restricted by regulatory authorities. However, no method was presently available for its specific detection. The large similarity in amino-acid sequences of collagens from different species make their immunochemical differentiation difficult when using polyclonal antibodies raised against the whole molecule [A. Venien, D. Levieux, J. Immunoassay Immunochem., in press]. To obtain bovine-specific antibodies, we immunized rabbits against putative species-specific sequences of the bovine collagen alpha 1(I) chain. Using these antibodies, an indirect ELISA was developed to allow a quick and easy differentiation between bovine and porcine gelatines. Moreover, a competitive indirect ELISA was found suitable to detect bovine gelatine in porcine gelatine purchased from laboratory chemicals suppliers down to a dilution of 2-4 parts per 1000 with CVs ranging from 5.7 to 7.7%. When testing mixtures of the largest possible range of industrial batches of bovine and porcine gelatines (skin/hides or bones origin, acid or alkaline processes, high or low Bloom) the detection limit was down to a dilution of 8 parts per 100 bovine gelatine in porcine gelatine. These ELISAs could be routinely used by pharmaceutical and food manufacturers to secure their supply chain.
Venien
A
A
INRA, Theix, Immunochemistry Unit, Meat Research Station, 63122 Saint-Genès-Champanelle, France.
Levieux
D
D
D000818:Animals; D000906:Antibodies / Q000737:chemistry; D001667:Binding, Competitive; D002417:Cattle; D003094:Collagen / Q000737:chemistry; D024042:Collagen Type I / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D004345:Drug Industry / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005780:Gelatin / Q000737:chemistry / Q000494:pharmacology*; D010455:Peptides / Q000737:chemistry; D013045:Species Specificity; D013552:Swine; D014443:Tyrosine / Q000737:chemistry
D000906:Antibodies; D024042:Collagen Type I; D010455:Peptides; D014443:Tyrosine; D005780:Gelatin; D003094:Collagen