An ELISA optimized for porcine epidemic diarrhoea virus detection in faeces

Abstract

Monoclonal antibodies to porcine epidemic diarrhoea virus (PEDV) membrane protein M were prepared and used for the comparative assessment of three blocking ELISA variants to detect PEDV. The competitive blocking ELISA (CB-ELISA) format showed the highest sensitivity, allowing detection of 10(2.5) plaque-forming units of PEDV/ml in culture medium. Its specificity was verified by inclusion of control samples containing transmissible gastroenteritis virus (TGEV) and rotavirus A in each analysis. Eighty porcine field samples of faeces obtained from 38 herds affected with diarrhoea were examined, and PEDV was found in 15 (19%) samples from 6 (16%) herds. The suitability of the CB-ELISA for the screening herds in epizootiologic situations is discussed.

Fig. 1

Western blot analysis of PEDV antibodies. After separation in 12% polyacrylamide gel, purified PEDV and a low molecular weight standard (LMW) were transferred to a nitrocellulose membrane. A part of the membrane with the LMW standard was stained with colloid gold (lane 1). The other lanes with PEDV were incubated with swine antiserum to PEDV (lane 2), mAbPEDV 3C8/2H6 (lane 4), C2/D5 (lane 5), 2B7 (lane 6), or with diluting solution alone (lanes 3 and 7). After incubation with peroxidase conjugates to swine (lanes 2 and 3) and mouse (lanes 4–7) immunoglobulins, the reaction was visualised by incubation in a solution containing the chromogen DAB.