Collaborative trial for validation of a real-time reverse transcriptase-polymerase chain reaction assay for detection of central nervous system tissues as bovine spongiform encephalopathy risk material: part 1.
A collaborative trial was conducted to evaluate a real-time reverse transcriptase-polymerase chain reaction (RT-PCR) assay for detection of central nervous system (CNS) tissues in meat products (e.g., sausages). The method is based on the detection of ruminant glial fibrillary acidic protein (GFAP) mRNA by applying real-time RT-PCR. The assay was evaluated through a multicenter trial involving 12 participating laboratories that received coded cDNA obtained from 3 different types of sausages. The participants used 5 different real-time detection systems. The results obtained in this validation revealed that this real-time RT-PCR assay performed well in the different laboratories with a detection limit of at least 0.1% CNS in those test materials that contained strongly heat-treated samples (sausages cooked at 120 degrees C) and the medium heat-treated samples (sausages cooked at 80 degrees C). The detection limit of liver sausages was determined to be 0.2% of CNS. Neither the samples with no CNS additive nor the bovine DNA and the negative control containing 100% swine brain gave any positive signals. The presented results indicate that the real-time RT-PCR assay was just as reproducible between laboratories, as repeatable within a laboratory, could reliably be used for detection of bovine spongiform encephalopathy risk material in meat and meat products, and signify that it may be used with confidence in any laboratory.
Abdulmawjood
Amir
A
Institute of Veterinary Food Science, Frankfurter Str, Giessen, Germany. amir.abdulmawjood@vetmed.uni-giessen.de
Schönenbrücher
Holger
H
Bülte
Michael
M
D000818:Animals; D002417:Cattle; D002490:Central Nervous System / Q000737:chemistry*; D003299:Cooperative Behavior; D016643:Encephalopathy, Bovine Spongiform / Q000635:transmission; D005506:Food Contamination / Q000032:analysis*; D005904:Glial Fibrillary Acidic Protein / Q000032:analysis* / Q000235:genetics*; D006358:Hot Temperature; D008461:Meat Products / Q000032:analysis*; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods* / Q000706:statistics & numerical data; D012307:Risk Factors
D005904:Glial Fibrillary Acidic Protein; D012333:RNA, Messenger