A monoclonal antibody‐based antigen capture enzyme‐linked immunosorbent assay for identification of infectious bronchitis virus serotypes

Abstract

SUMMARYAn antigen-capture enzyme-linked immunosorbent assay (C-ELISA) was developed for detection and identification of infectious bronchitis virus (IBV) serotypes Arkansas, Connecticut, and Massachusetts using monoclonal antibodies (MAbs) specific to the S1 glycoprotein of the respective serotype. The assay (designed as a doubleantibody sandwich assay) gave the best results when the S1-specific MAb, antigen, and chicken serum were of the same serotype. However, when a group-specific (M glycoprotein-specific) MAb was used for antigen capture, a distinctive pattern of cross-reactivity was observed between the antigens and heterologous chicken sera, suggesting a complex distribution of epitopes on the IBV M glycoproteins. Treatment of antigen with NP40 enhanced the ELISA signal only when the M glycoprotein-specific MAb was used for antigen capture. Although C-ELISA was inconsistent in detecting IBV in chicken tissue homogenates, it was highly effective in detecting the virus in allantoic fluid after the homogenates were given one chicken embryo passage.

Detection of infectious bronchitis virus

“…Because of these relatively high amounts of virus that are needed for detection by antigen ELISA, the sensitivity for detecting IBV antigen directly in chicken organs is low. Naqi et al (1993) failed to detect IBV by ELISA in tracheas of susceptible chickens after inoculation, while Nagano et al (1990) reported that the ELISA detected IBV for only 2 days, while virus was isolated for at least 7 days (the end of the experiment). Ignjatovic & Ashton (1996) could not detect antigen by ELISA after infection of chickens with a commercial vaccine, but did isolate the virus.…”

Detection of Infectious Bronchitis Virus and Specific Anti- Viral Antibodies Using a Concanavalin A–Sandwich–ELISA

“…Con A-S-ELISA did not detect IBV antigens in trachea and lung homogenates easily, but it was highly efficient in detecting this virus after one embryo passage, depicting a general performance not statistically different from the VI test and showing a high reproducibility (Table 1). Furthermore, the Con A-S-ELISA must be considered as sensitive and specific as an S-ELISA applied for the detection of IBV antigen by Naqi et al (21), using a set of mAbs against the different IBV proteins, as capture antibodies.…”

“…Enzyme Linked Immunosorbent Assays (ELISAs) have been developed and proved their efficacy, either for the detection of IBV antigen (1,12,15,19,21,34), or anti-IBV antibodies (2,3,4,8,16,22,29).…”

“…Alternatively, polyclonal or monoclonal anti-IBV capture antibodies have been adsorbed on the solid phase in sandwich-ELISA methods for trapping these viruses (12,19,21). Whole virus purification requires propagating large quantities of virus in eukaryotic systems and whereas the most reliable serodiagnostic reagents require highly purified antigen, purification of IBV with its highly glycosylated spike protein is difficult and expensive (22).…”

Reverse Transcription Loop-Mediated Isothermal Amplification for the Rapid Detection of Infectious Bronchitis Virus

“…Immunoassays use IBV-specific monoclonal antibodies to detect the virus in direct or indirect fluorescent antibody and enzyme-linked immunosorbent assay formats. Although more rapid and simpler than virus isolation, immunoassays tend to lack specificity and sensitivity to some extent and can not detect all strains of IBV (Karaca and Naqi, 1993;Karaca et al, 1992;Naqi et al, 1993). Molecular assays for the detection of IBV are used commonly because they provide highly specific and sensitive results and detect viral RNA directly from clinical samples or from virus isolated in a laboratory host system.…”