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: The food industry must provide consumers with products which are innocuous, genuine, desirable and nutritive. To assure consumers that food has these characteristics, it is convenient to have appropriate controls of their hygiene state and to detect fraudulent practices, both of which are possible using adequate analytical techniques. However, the analysis of food has its own limitations such as specificity, reproducibility, sensitivity, economy, ease of use and standardization. Immunological techniques have objective advantages which merit their development and further use in the food industry. It has been widely demonstrated that immunological methods can be used in the food industry as reliable analytical techniques. Antibody‐based analytical methods for determining meat species and detecting adulteration of milk have been developed. We review the efforts made to develop immunological reactions and techniques to detect fraudulent replacements of foods. The selection of antigens as immunogens, the development of various polyclonal and monoclonal antibodies, and the selection of immunological techniques from immunodiffusion and immunoelectrophoresis to ELISA are described. The development of recombinant antibodies as an alternative for replacement or substitution of previously developed polyclonal or monoclonal antibody reagents will also be considered.Food and Agricultural Immunology 01/1994; 6(1):95-104. DOI:10.1080/09540109409354817 · 0.98 Impact Factor
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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 11/1994; 66(3):411 - 415. DOI:10.1002/jsfa.2740660321 · 1.88 Impact Factor