Indirect ELISA for detection of goats' milk in ewes' milk and cheese

An indirect ELISA (enzyme-linked immunosorbent assay) has been developed successfully for the detection of defined amounts of goats' milk (1–25%) in ewes' milk and cheese. The assay uses polyclonal antibodies raised in rabbits against goats' caseins (GC). The anti-GC antibodies were recovered from the crude antiserum by immunoadsorption and elution from a column containing immobilized goats' caseins. The anti-GC antibodies were biotinylated and rendered goats' milk specific by mixing them with lyophilized bovine and ovine caseins. The assay was developed in a non-competitive ELISA format and it comprised coating plates with extracts from samples. ExtrAvidin-peroxidase was used to detect the biotinylated anti-GC antibodies bound to goats' caseins immobilized on 96-well plates. The colour developed by the subsequent enzymic conversion of the substrate resulted in discernible differences in optical densities when assaying mixtures of ewes' milk and cheese containing variable amounts of goats' milk.