Production of a sensitive immunoassay for detection of ruminant and porcine proteins, heated to > 130°C at 2.7 bar, in compound animal feedstuffs

Sandwich ELISAs have been produced capable of detecting 1250 ppm ruminant and porcine heat stable proteins, which have been heated to 130 degreesC, 2.7 bar (absolute pressure), for 30 min and incorporated into compound animal feedstuffs (CAFs). Removal of most of the proteins derived from CAFs and rendered animal material by ammonium sulphate precipitation, followed by concentration of the specific heat stable proteins for species testing, proved to be of considerable benefit in amplifying test sensitivity (GB Patent 9317199.9, 18 August 1993; World Patent PCT/GB94/01715, 4 August 1994). Increased sensitivity was also achieved by both pre-heating the sample prior to protein extraction to overcome problems attributed to the presence of oils or fats, and optimising ELISA incubation stages. Occasional false positives, resulting from non-specific binding of lectins in plant material such as salseed, are identified through the use of a separate ELISA utilising non-immune sera.