Development of a unique sandwich enzyme-linked immunosorbent assay based on monoclonal antibodies for the specific detection of the egg drop syndrome virus

ABSTRACT

An abrupt decrease in egg production and quality in laying hens can be brought on by the avian adenovirus known as the egg drop syndrome virus (EDSV). So far, there are no efficient commercial diagnostic approaches for EDSV. In the present study, we first purified two monoclonal antibodies (mAbs 5G4 and 6G6) specifically against the EDSV fibre protein. The mAb 5G4 was then employed as capture antibody. Additionally, the detection antibody 6G6 (HRP-6G6) was conjugated with horseradish peroxidase. Consequently, based on these two mAbs, we developed a sandwich enzyme-linked immunosorbent assay (ELISA) for EDSV detection. Specificity evaluation demonstrated that only EDSV gave a positive result while other avian viruses showed negative results. Sensitivity investigation showed that the limit of detection of EDSV was 10^2.9 TCID₅₀/ml and the limit of detection of the purified His-Fiber was 5 ng/ml. Furthermore, the screening of samples from infected chickens showed that this method was very effective and could be applied to the detection of EDSV in infected samples. In summary, the sandwich enzyme-linked immunosorbent assay developed here provides an efficient, sensitive, and large-scale method for the specific diagnosis of EDSV.

RESEARCH HIGHLIGHTS

• A sandwich ELISA was developed to detect EDSV using the mAbs 5G4 and HRP-6G6.

• The sandwich ELISA maintained high specificity and sensitivity.

• The sandwich ELISA had equivalent consistency with real-time PCR assay.

KEYWORDS:

• Egg drop syndrome virus
• sandwich enzyme-linked immunosorbent assay
• fibre protein
• monoclonal antibody
• detection of antigen
• limit of detection

Ethical statement

The bird experiments were approved by the Key Laboratory of Animal Immunology, Henan Academy of Agricultural Sciences, China, in line with its policies and procedures.

Disclosure statement

No potential conflict of interest was reported by the authors.

Additional information

Funding

This work was supported by grants from the National Natural Science Funds of China [grant number 32002284], the Key Scientific and Technological Research Projects of Henan Province [grant number 232102110105], and Henan Academy of Agricultural Sciences [grant number 2022YQ22].