COMPARATIVE ANALYSIS OF METHODS FOR OBTAINING ANTIGENS FOR THE DIAGNOSIS OF EQUINE RHINOPNEUMONITIS

Abstract

Methods for obtaining antigens for the diagnosis of equine rhinopneumonitis, also known as viral abortion of horses, using enzyme-linked immunosorbent assay were studied. The methods were based on stepwise centrifugation and precipitation with polyethylene glycol. The vaccine strain CB/69 of equine herpesvirus type 1, cultivated in SPEV cell culture, was used as the source material. The infectious titer of the virus was 6.5 lg TCD₅₀/ml. It was found that the most purified antigen was obtained from free virions by stepwise centrifugation followed by purification in a sucrose density gradient. However, this method requires considerable time, material resources, and specialized equipment, and is characterized by a low yield: 400 µl of antigen was obtained from 105 ml of the initial virus-containing suspension. The antigen isolated from the cell-associated viral fraction had a higher yield of 650 µl but contained more cellular impurities. Polyethylene glycol precipitation was technologically simpler and provided a higher antigen yield of 1.5 ml from 125 ml of suspension; however, the resulting preparation was unsuitable for ELISA, despite being effective in the hemagglutination inhibition test. The results indicate that the method for obtaining EHV-1 antigen should be selected depending on the intended diagnostic application. These findings may be used in the development of methodological recommendations for obtaining EHV-1 antigens for the diagnosis of equine rhinopneumonitis.