title: A new homogeneous assay for high throughput serological diagnosis of brucellosis in ruminants.
abstract: The control and eradication of brucellosis is highly desirable but heavily resource intensive as high throughput serological testing is required. The aim of this study was to meet the needs of high throughput screening laboratories involved in this process through the development of a new assay. An existing cELISA used for the serodiagnosis of brucellosis in ruminants was converted to an AlphaLISA homogenous proximity based assay. This assay requires no separation steps and can be performed in low volume microtitre format. The Brucella AlphaLISA was validated on a panel of bovine, ovine and caprine sera from infected and uninfected animals. The diagnostic sensitivities (>96%) and specificities (>98%) obtained compared well to those from cELISA, iELISA and FPA performed on the same samples. The AlphaLISA met the testing criteria set for ELISAs as defined by the OIEELISA standards and had an analytical sensitivity similar to that of the parent cELISA. The method was also used on a small panel of serum samples from cattle that were experimentally infected with Yersinia enterocolitica O:9. Some false positive reactions were obtained as was also the case with results from FPA, iELISA, cELISA, CFT and SAT. Despite this, the methodological advantages of the AlphaLISA mean that this assay is well suited to high throughput serodiagnosis. This report is the first description of the use of AlphaLISA to detect pathogen specific antibodies. Furthermore, the relative ease with which the cELISA was converted to this platform indicates that this technology is ready to meet the high throughput testing requirements for the diagnosis of many other diseases.
authors.affiliation: Brucella Research Group, Statutory and Exotic Bacteria, Veterinary Laboratories Agency Weybridge, New Haw, Addlestone, Surrey, UK. j.mcgiven@vla.defra.gsi.gov.uk
authors.forename: John A
authors.initials: JA
authors.lastname: McGiven
authors.forename: Jason
authors.initials: J
authors.lastname: Sawyer
authors.forename: Lorraine L
authors.initials: LL
authors.lastname: Perrett
authors.forename: Simon D
authors.initials: SD
authors.lastname: Brew
authors.forename: Nicola J
authors.initials: NJ
authors.lastname: Commander
authors.forename: Alan
authors.initials: A
authors.lastname: Fisher
authors.forename: Stuart
authors.initials: S
authors.lastname: McLarnon
authors.forename: Kate
authors.initials: K
authors.lastname: Harper
authors.forename: Judy A
authors.initials: JA
authors.lastname: Stack
mesh_terms: D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood*; D000918:Antibody Specificity; D002003:Brucella abortus / Q000276:immunology*; D002006:Brucellosis / Q000175:diagnosis* / Q000382:microbiology / Q000662:veterinary; D002007:Brucellosis, Bovine / Q000175:diagnosis* / Q000382:microbiology; D002417:Cattle; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay / Q000662:veterinary*; D005189:False Positive Reactions; D015511:Goat Diseases / Q000175:diagnosis* / Q000382:microbiology; D006041:Goats; D011237:Predictive Value of Tests; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012698:Serologic Tests / Q000662:veterinary*; D012756:Sheep; D012757:Sheep Diseases / Q000175:diagnosis* / Q000382:microbiology; D015008:Yersinia enterocolitica / Q000276:immunology
chemical_list: D000907:Antibodies, Bacterial