Partial purification of horse‐specific soluble muscle proteins by immunoadsorption chromatography
ABSTRACT Immunoadsorption chromatography has been used to isolate horse-specific soluble muscle proteins from crude horse protein extracts. Horse-specific polyclonal antibodies against soluble horse muscle proteins immobilised on a Protein A-Sepharose CL-4B matrix were used to adsorb the corresponding antigens. Analysis of the crude soluble horse muscle proteins by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) showed the existence of 20 protein subunits in the gel. SDS-PAGE of the proteins recovered by affinity chromatography showed nine protein subunits, three of which were enriched after the immunopurification step. These affinity-recovered horse-specific soluble muscle proteins may be used as antigens in the development of monoclonal antibodies to detect and quantify horse meat in raw, unheated meat mixtures.
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ABSTRACT: A stable hybridoma cell line (DD3) has been produced secreting a monoclonal antibody specific for horse muscle proteins. The DD3 monoclonal antibody (mAb) did not show significant cross-reactivity when tested against beef, chicken, pig and soya proteins or bovine caseins, gelatine and bovine serum albumin. The DD3 mAb was used in an indirect ELISA format for the detection of defined amounts of horse meat (10–500 g kg-1) in beef meat mixtures. Immunorecognition of monoclonal antibodies adsorbed to horse meat adsorbed onto the ELISA plate was made with rabbit anti-mouse immunoglobulins conjugated to the enzyme horseradish peroxidase. Subsequent enzymic conversion of the substrate gave clear optical density differences when assaying mixtures of minced beef containing different amounts of horse meat.Journal of the Science of Food and Agriculture 09/2006; 66(3):411 - 415. · 1.76 Impact Factor