Development of solid phase competitive enzyme-linked immunosorbent assay to detect foot-and-mouth disease virus A serotype specific antibodies based on immunoglobulin Y antibody.

Abstract

The aim of this study was to using purified chicken antibody (IgY) for developing solid phase competitive (SPC) enzyme-linked immunosorbent assay (ELISA) to detect the foot-and-mouth disease virus (FMDV) A serotype. After immunization of chickens, polyclonal immunoglobulin (IgY) antibodies were extracted and purified from egg yolk and yield was about 5.00 mg mL^-1 of yolk as well as near 0.40 mg mL^-1 of specific IgY antibody against FMDV serotype A. Also, optimized sucrose density gradient method produced 228 µg mL^-1 whole virus which is much higher than that of the conventional method of sucrose density gradient method. The optimum concentration of purified capture IgY and bind type A antigen were 0.50 µg and 0.10 µg per well, respectively. The OD values < 0.70 were considered positive, and values ≥ 0.70 were negative for in-house kit base on standard controls. Statistical analysis base on 80 serum samples showed the 96.66% sensitivity, 100% specificity, 100% positive predictive value, 90.90% negative predictive value, 97.50% accuracy, and 98.33% reliability for serum samples for two commercial and in-house kits. The SPCE developed based on IgY antibody is a suitable alternative for the detection of antibodies after vaccination against type A FMDV with high sensitivity and specificity. The present research demonstrated the possibility of commercial development of the SPCE kit using IgY antibodies for the detection of FMDV antibodies in serum samples with adequate sensitivity and accuracy.