Comparison of native and subunit antigens as ELISA reagents for the detection of antibodies against scabby mouth virus.
A simple ELISA test to detect antibodies against scabby mouth virus (SMV) has been developed. Native whole virions and subunits of SMV generated by boiling the virus in the presence of sodium dodecyl sulphate (SDS) detergent and beta-mercaptoethanol were compared as ELISA assay reagents using naive and hyperimmune sera from sheep and rabbits. Approximately 2 x 10(4) intact virus particles per microtiter well were required to generate a positive to negative signal of 0.8:0.3 ELISA O.D. units when the serum was used at a dilution of 1/100. In contrast, total subunit antigen generated by disrupting and coupling of 250-500 virions per well provided a signal ratio of 1.58:0.3 ELISA O.D. units at a serum dilution of 1/250. Total subunit antigens were therefore 400 times more economical to use than intact virions. In addition, subunit antigens could be readily bound to microtiter plates without the need for removal of the SDS. Secondly, it was not necessary to block non-specific binding sites on the plate with blockers such as gelatin and skim-milk, thereby shortening the time needed to complete the ELISA assay. The total subunit antigen ELISA test was used to detect seroconversion in new born lambs where there was an occurrence of SMV infection in housed sheep. Three bleeds were taken at fortnightly intervals and the ELISA results showed that 9 out of 15 lambs were seropositive for all bleed points. Four of the lambs showed a sequential rise in titer while only one lamb failed to seroconvert.(ABSTRACT TRUNCATED AT 250 WORDS)
Chin
J C
JC
Elizabeth Macarthur Agricultural Institute, NSW Agriculture, Sydney, Australia.
Petersen
R K
RK
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral; D002478:Cells, Cultured; D004474:Ecthyma, Contagious / Q000175:diagnosis* / Q000276:immunology*; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000662:veterinary*; D015151:Immunoblotting / Q000379:methods; D008297:Male; D008623:Mercaptoethanol; D009923:Orf virus / Q000254:growth & development / Q000276:immunology / Q000302:isolation & purification*; D011817:Rabbits; D012756:Sheep; D012967:Sodium Dodecyl Sulfate; D013737:Testis; D015164:Vaccines, Inactivated; D014765:Viral Vaccines; D014771:Virion / Q000302:isolation & purification
D000914:Antibodies, Viral; D000956:Antigens, Viral; D015164:Vaccines, Inactivated; D014765:Viral Vaccines; D012967:Sodium Dodecyl Sulfate; D008623:Mercaptoethanol