The development and application of an indirect Elisa test for the detection of antibodies to bovine respiratory syncytial virus in blood serum

Abstract

ABSTRACT:We developed an indirect enzyme-linked immunosorbent assay (ELISA) for detection of serum antibodies to bovine respiratory syncytial virus. For evaluation of the newly developed ELISA, field sera collected from 549 head of cattle in the Czech Republic were tested in parallel by a serum neutralization test. The tests showed 98.36% agreement. The specificity and sensitivity of the ELISA relative to serum neutralization test was 97.00% (226/233) and 99.37% (314/316), respectively. Tissue culture-grown viral antigen was used in the tests. The corrected optical density (COD) of each sample tested at dilution 1/100 was expressed as a percentage of the COD of a positive reference serum included on each plate, this value was the sample/positive (S/P) ratio. We determined the relationship between the S/P ratio (%) obtained at a dilution 1/100 and the end point titer calculated by serum neutralization test (r = 0.9743). The ELISA test was evaluated by testing acute and convalescent (3 wk later) serum pairs from 9 head of cattle with confirmed BRSV infection for demonstration of seroconversion. The ELISA test demonstrated a clear increase of the S/P ratio (%) between acute and convalescent serum pairs (on average 42.2 ± 13.1).Keywords: bovine respiratory syncytial virus (BRSV); respiratory tract pathogen; cattle; ELISA A list of abbreviations: BRSV bovine respiratory syncytial virus, SN test serum neutralization test, HIT haemagglutination inhibition test, FAT fluorescent antibody test, CPE cytopathic effect, OD optical density, PBS-T phosphate buffered saline with Tween 20, MEM minimal essential medium, RT-PCR reverse transcription-polymerase chain reaction.

Expression and Purification of Human Arp1 Protein and Rapid Preparation of Polyclonal Antibody

“…[15][16][17][18][19] The purified antigen, which was dissolved to the concentration of 10 mg=mL in 50 mM carbonate buffer (pH 9.6), was coated on immunoplates at 100 mL per well, kept at 37 C for 2 hr, and then at 4 C for 48 hr. Serial twofold dilutions of antibodies from 200-fold to 102,400-fold were used for reactions with the MBP-FD antigen on the immunoplates.…”