Use of monoclonal antibodies in blocking ELISA detection of transmissible gastroenteritis virus in faeces of piglets

Abstract

Monoclonal antibodies (mAb) to the transmissible gastroenteritis virus (TGEV) nucleoprotein (N) and membrane protein (M) were prepared and used for the comparative assessment of three blocking ELISA variants to detect TGEV. The competitive blocking ELISA format showed the highest sensitivity, allowing detection of 10(3) TCID50 TGEV/ml in culture medium. Ninety-nine porcine field faecal samples obtained from 37 herds affected with diarrhoea were examined, and various TGEV levels were found in nine samples from six herds. However, only in three samples were significant TGEV concentrations demonstrated. The relationship between incidence of TGEV gastroenteritis and the spread of porcine respiratory coronavirus infection in pig farms is discussed.

Figure 1

Western blot analysis of TGEV antibodies. After separation in 12% polyacrylamide gel, purified TGEV 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 colloidal gold (lane 1). The other lanes with TGEV were incubated with swine antisera to TGEV (lanes 2 and 3), mAbTGEV D3/G6 (lane 5), D7/G7 (lane 6), B7/F7 (lane 7) or with diluting solution alone (lanes 4 and 8). After incubation with peroxidase conjugates to swine (lanes 2–4) and mouse (lanes 5–8) immunoglobulins, the reaction was visualised by incubation in a substrate solution containing chromogen DAB. The localization of TGEV antigens S, N and M is indicated.

Figure 2

Detection of TGEV in infected cells by immunoperoxidase test. Direct immunoperoxidase detection of TGEV in a cell line PK‐15 12 hpi using the conjugate HRPO‐mAbTGEV D7/G7. Intensive staining of the cytoplasm proves propagation of TGEV in infected cells.