The reactions of monoclonal antibodies with structural proteins of mumps virus.

Abstract

Mouse hybridomas producing antibodies against structural proteins of mumps virus were established by fusion of FO or SP 2/0 myeloma cells with spleen cells from BALB/c mice immunized with purified preparations of egg-grown mumps virus. Ascites fluids collected after i.p. inoculation of mice were characterized by different serologic tests. By immune precipitation tests with [35S]methionine-labeled mumps virus polypeptides, 17 clones were found to produce antibodies against the nucleocapsid protein (NP), 11 against the polymerase (P) protein, 10 against the membrane (M) protein, 12 against the fusion (F) protein, and 24 against the hemagglutinin-neuraminidase (HN) protein. Competitive binding enzyme-linked immunosorbent assay (ELISA) tests were performed to determine the reactivity of the monoclonal antibodies with different antigenic sites of each structural component. The monoclonal antibodies directed against the NP, P, and M proteins identified a minimum of 10, 10, and 9 separate antigenic sites, respectively. The 12 clones directed against F were directed against a minimum of eight separate antigenic determinants. These antibodies did not neutralize the infectivity of the virus either in the absence or presence of anti-gamma-globulin. Only low capacity to block hemolysis (HL) activity of the virus was detected in clones directed against three of the eight antigenic sites. Based on their serologic reactivity, the 24 clones directed against the HN protein could be divided into four groups. The first group of clones could not inhibit any biologic activity of the protein. The second group consisted of two clones that blocked HL but did not block hemagglutination (HA) or neuraminidase (NA) activity. The third group, which included five clones, blocked HA, NA, and HL activity of the virus and had high neutralizing capacity. These clones were directed against three distinct antigenic sites. Two of the clones directed against one antigenic site could block NA activity only when a large substrate, fetuin, was used, but not when a small substrate, neuraminlactose, was used in the test. The fourth group included five clones that could block NA but not HA activity of the virus. These clones could neutralize the infectivity of the virus and had high capacity to block HL activity. In blocking experiments, all these antibodies reacted with one antigenic site. The reaction of all clones was tested in ELISA with four different strains of mumps virus. Each strain had unique antigenic sites. Variations were found in four, three, and three different antigenic sites of the NP, P, and HN proteins, respectively.