The effects of monoclonal antibodies on biologic activities of structural proteins of Sendai virus.

Abstract

Abstract Mouse hybridoma antibodies against structural proteins of Sendai virus were produced by fusion of FO myeloma cells with spleen cells from BALB/c mice immunized with purified preparations of egg-grown Sendai virus. Ascites fluids collected after i.p. inoculation of mice with 133 hybridoma cell lines were characterized by different serologic tests. By immune precipitation tests with 35S-methionine-labeled extracellular Sendai virions and cell-associated virus polypeptides, 10 clones were found to produce antibodies against the nucleocapsid protein (NP), 6 against the polymerase (P) protein, 4 against the membrane (M) protein, 7 against the fusion (F) protein, and 52 against the hemagglutinin-neuraminidase (HN) protein. The specificity of the remaining clones could not be determined. Selected mouse hybridoma antibodies against the five different structural proteins of Sendai virus were used in immune fluorescence to characterize the appearance of viral antigens in infected Vero cell cultures. Trace amounts of all five antigens were detectable 7½ hr postinfection. Antibodies against NP, P, and M proteins stained cytoplasmic inclusions varying in size from small granules to more confluent masses. Late after infection the large inclusions were frequently found in a perinuclear position. In contrast, antibodies to envelope components, HN and F, gave a dense fine granular fluorescence evenly distributed in all of the cytoplasma. Based on their serologic reactivity, the seven clones directed against the F protein could be divided into two groups. One group of clones comprised antibodies against a minimum of three distinct antigenic sites and was found to block the hemolysis (HL) activity and infectivity after addition of anti-γ-globulin to the test, whereas the other group of two specific clones could not block these biologic activities. The 52 clones directed against the HN protein could be divided into four groups. The first group of clones could not inhibit any biologic activity in the absence of anti-y-globulin. The second group blocked hemolysis, but did not block hemagglutination (HA) or neuraminidase (NA) activity. The third group, which included six clones, blocked all biologic activities of the virus. These clones were directed against two distinct antigenic sites. The fourth group included seven clones, all of which only could block NA activity of the virus. In blocking experiments, all these antibodies reacted with one antigenic site. Passive immunization of mice with monoclonal antibodies against the F protein blocking hemolysis and the third and fourth group of monoclonal antibodies against the HN protein protected mice against experimental Sendai virus infection.