Characterization of immune response to synthetic peptides derived from the hemagglutinin of measles virus.

We describe here four synthetic peptides derived from the hemagglutinin of measles virus. The peptides were predicted by a computer program combining hydrophilicity, flexibility, surface probability, secondary structure and antigenic index parameters of the amino acid sequence of measles virus hemagglutinin. Rabbits were immunized with the synthesized peptides conjugated to purified protein derivative using immunostimulating complex as adjuvant. Anti-peptide antisera raised in rabbits against the peptide conjugates reacted well with the homologous peptides and with measles virus antigen as tested with plate ELISA. None of these sera had either neutralizing or hemagglutination inhibiting antibody or reacted with measles hemagglutinin protein in Western blot and reacted weakly in immunofluorescence. Human sera positive for measles virus antibody reacted with the synthesized peptides indicating that the selected locations function as partial antigenic sites.

Analysis of antibody response to synthetic peptides derived from the fusion protein of measles virus.

A computer program combining of hydrophilicity, flexibility, surface probability, secondary structure and antigenic index parameters of the amino acid sequence of measles virus (MV) fusion protein was used to select four possible epitopes. Rabbits were immunized with the synthesized peptides conjugated to purified protein derivative using the homobifunctional cross-linker bis-sulfosuccinimidyl suberate. Immune stimulating complexes were prepared with the peptides conjugated to the purified protein derivative carrier using a dialysis method. All antisera raised in rabbits against the peptide conjugates had a high titer to the homologous peptides and reacted well with denatured MV as tested by plate ELISA. None of the sera had neutralizing antibody. Human sera positive for MV antibody reacted strongly with the synthesized peptides indicating that the selected locations function as partial antigenic sites. Antisera against peptide conjugates reacted weakly in immunofluorescence and none of these antisera reacted with purified MV proteins in Western blot. The results obtained in this study indicated that although the computer program could not predict epitopes important for the neutralization of the MV, the predicted epitopes are useful for detecting antibodies against MV.

Generation of rubella virus-neutralising antibodies by vaccination with synthetic peptides.

Four short peptides from rubella virus proteins E1 and E2, predicted to contain B cell epitopes, were used to vaccinate BALB/c mice. Sera from peptide-vaccinated animals reacted with viral antigens in ELISA and three of the four induced virus-neutralising antibody (nAb) responses. Peptide PY4, in contrast to the others, induced IgG2a responses upon vaccination and stimulated spleen cells in vitro produced IFN gamma in the absence of IL-5. It was reasoned that vaccination with PY4 caused Th1 subset activation, the appropriate type of response for anti-viral immunity and hence the efficient neutralising antibody response. Presentation of peptide for vaccination proved to be as important as the sequence. Similar profiles of IgG1 and IgG2a were detected in the sera of mice vaccinated with PY4 in Freund's complete adjuvant or alum; however nAb responses were not found when alum was used.

Toxoplasma gondii: detection of antibodies in human saliva and serum.

Saliva samples from 27 patients with a recent toxoplasma infection were tested for specific IgG, IgM and IgA antibodies to Toxoplasma gondii. Thirteen of the 27 saliva samples were positive for IgG anti-T. gondii by direct agglutination and 8 of the 27 were positive for IgM anti-T. gondii by an immunosorbent agglutination assay. Twenty of the 27 saliva samples were positive for IgG antibody on toxoplasma immunoblots with three major immunodominant antigens; 38, 30 and 35 kDa. IgA results on toxoplasma immunoblots were positive for all three groups tested, recently infected patients, chronically infected and seronegative adults without distinguishing between them. The 35 and 43 kDa antigens were the most frequently detected proteins. IgM in saliva gave negative or very weak reactions. None of the eight seronegative or the 17 chronically infected adults gave positive results in any of the tests performed to detect IgG or IgM in saliva. Serial saliva and serum samples from a laboratory-infected patient were collected and tested for toxoplasma-specific IgG, IgM and IgA. IgG in saliva was detected by 27 days post infection (p.i.) and was negative by 81 days p.i.; it detected mainly the 38 and 30 kDa antigens. IgM in saliva was detected by 11 days p.i. and was negative by 81 days p.i., with no reaction on immunoblots.

Preparation and properties of Vibrio cholerae antifimbrial antibody.

The presence of fimbriae on the Vibrio cholerae strains used was assessed by pellicle formation, haemagglutination activity and electron microscopy. Fimbrial suspensions were prepared by shearing them off the organisms, then separating them from other components by absorbing them on to rabbit red blood cells. Rabbits were then immunized with the fimbrial-red cell suspensions and the antibodies evoked were titrated by haemagglutination inhibition, agglutination, vibriocidal and immobilization techniques.

Reactions of extracts of Reiter treponeme with syphilitic and non-syphilitic human sera in the single radial haemolysis technique.

A single radial haemolysis (SRH) technique, using Reiter protein or Reiter lysate as the coating antigen, was investigated. Results obtained with syphilitic and presumed non-syphilitic human sera were compared with results obtained in the absorbed fluorescent treponemal antibody test (FTA-ABS), the Reiter protein complement fixation test ( RPCFT ), the Venereal Diseases Research Laboratory Slide test (VDRL) and the Cardiolipin Wasserman reaction ( CWR ). The SRH reaction, with either Reiter antigen, was more sensitive than any of the screening tests ( RPCFT , VDRL and CWR ) for detecting positive syphilitic antibodies. Although the SRH test used almost the same materials as the RPCFT , it was appreciably more sensitive for the detection of the group-specific antibodies in syphilitic human serum.

A proposed life cycle for the Reiter treponeme.

Direct microscopical observations of single developing cysts in sealed slide microcultures prepared from 5-days-old tube cultures of the Reiter treponeme revealed two distinct phases in the life cycle. In one phase transverse fission was the main method of multiplication while in a second phase, occurring when conditions in the medium became unfavourable for propagation, cysts developed. These could release large numbers of actively motile treponemes when returned to optimum growth conditions. These observations, together with results of the dilution method for the calculation of the Most Probable Number and the absence of a response in treponemes killed by high temperature (45 degrees C) or abnormal pH (10), showed that the cysts were viable and a mode of propagation for the Reiter treponeme.

Reiter haemagglutination test: a screening test for syphilis.

Using an ultrasonicate of the Reiter treponeme as antigen the Reiter haemagglutination test (RHA) was evaluated as a serological test for syphilis. Comparison of the results of the cardiolipin Wassermann reaction, Reiter protein complement-fixation test, the fluorescent treponemal antibody-absorbed (FTA-ABS) test, the Treponema pallidum haemagglutination test (TPHA) (at dilutions of 1/16 and 1/80), and the Venereal Disease Research Laboratory test with those of the RHA showed that the RHA was sensitive (85.8%) and agreed well (85.8%) with the FTA-ABS test result. Simplicity, sensitivity, availability of the antigen, and the very low cost of this test support its use as a first-line screening test for syphilis.

Some reactions of influenza viruses adsorbed to polystyrene for enzyme immunoassay.

Three influenza viruses adsorbed to the surface of polystyrene tubes exhibited the property of haemadsorption, but only two of the neuraminidases were still active. Reactions of the viruses with rabbit antisera indicated that at least some of the virus particles were ruptured by adsorption to the polystyrene and the exposed nucleoprotein could react with antibody. Enzyme immunoassays involving direct adsorption of influenza virus particles to polystyrene are unlikely, therefore, to differentiate between strains of influenza A.