A bioinformatics approach to the development of immunoassays for specified risk material in canned meat products.
A bioinformatics approach to developing antibodies to specific proteins has been evaluated for the production of antibodies to heat-processed specified risk tissues from ruminants (brain and eye tissue). The approach involved the identification of proteins specific to ruminant tissues by interrogation of the annotation fields within the Swissprot database. These protein sequences were then interrogated for peptide sequences that were unique to the protein. Peptides were selected that met these criteria as close as possible and that were also theoretically resistant to either pepsin or trypsin. The selected peptides were synthesised and used as immunogens to raise monoclonal antibodies. Antibodies specific for the synthetic peptides were raised to half of the selected peptides. These antibodies have each been incorporated into a competitive enzyme-linked immunosorbent assay (ELISA) and shown to be able to detect the heat-processed parent protein after digestion with either pepsin or trypsin. One antibody, specific for alpha crystallin peptide (from bovine eye tissue), was able to detect the peptide in canned meat products spiked with 10% eye tissue. These results, although preliminary in nature, show that bioinformatics in conjunction with enzyme digestion can be used to develop ELISA for proteins in high-temperature processed foods and demonstrate that the approach is worth further study.
Reece
Paul
P
Food and Environment Research Agency, York, YO41 1LZ, UK. p.reece@csl.gov.uk
Bremer
Monique
M
Stones
Robert
R
Danks
Christopher
C
Baumgartner
Sabine
S
Tomkies
Victoria
V
Hemetsberger
Claudia
C
Smits
Nathalie
N
Lubbe
Walter
W
D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology; D000918:Antibody Specificity; D000937:Antigen-Antibody Reactions; D002417:Cattle; D019295:Computational Biology; D016643:Encephalopathy, Bovine Spongiform / Q000517:prevention & control / Q000635:transmission; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D005506:Food Contamination / Q000032:analysis* / Q000517:prevention & control; D005519:Food Preservation; D006041:Goats; D008460:Meat / Q000032:analysis*; D008461:Meat Products / Q000032:analysis*; D051379:Mice; D008807:Mice, Inbred BALB C; D010434:Pepsin A / Q000378:metabolism; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry / Q000276:immunology; D011506:Proteins / Q000032:analysis* / Q000276:immunology; D015203:Reproducibility of Results; D012307:Risk Factors; D012680:Sensitivity and Specificity; D012756:Sheep; D012984:Software; D013552:Swine; D013696:Temperature; D014357:Trypsin / Q000378:metabolism
D000911:Antibodies, Monoclonal; D010455:Peptides; D011506:Proteins; D014357:Trypsin; D010434:Pepsin A