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. 1984 Jan;43(1):421–428. doi: 10.1128/iai.43.1.421-428.1984

Monoclonal antibodies distinguish phase variants of Coxiella burnetii.

J C Williams, M R Johnston, M G Peacock, L A Thomas, S Stewart, J L Portis
PMCID: PMC263449  PMID: 6418662

Abstract

Monoclonal antibodies (MAbs) directed against phase I and II variants of Coxiella burnetii were produced by fusing myeloma SP2/O-AG 14 cells with spleen cells from mice immunized with the chloroform-methanol extraction residue of phase I whole cells. Two hybridoma clones which distinguished the phase variants by microimmunofluorescence assay were isolated and characterized. The MAbs showing specificity for phase I cells (MAbI-1, immunoglobulin G, subclass 3 kappa) reacted with the hot phenol-water extract of phase I C. burnetii in immunodiffusion and enzyme-linked immunosorbent assays. MAbI-1 reacted with high-molecular-weight components from phase I phenol-water extract and whole cell in an immunoblot assay. Specificity of MAbI-1 for a carbohydrate epitope in the phenol-water extract was demonstrated by periodic acid inactivation of binding by a competitive enzyme-linked immunosorbent assay. Phase I antigenic sites were apparently well represented on the surface of cells as demonstrated by complete fluorescence and microagglutination. The MAb showing specificity for phase II cells (MAbII-1, immunoglobulin G, subclass 2b kappa) reacted with whole cells in the microimmunofluorescence assay, microagglutination test, complement fixation test, and the enzyme-linked immunosorbent assay. MAbII-1 reacted specifically with a 29,500-dalton surface protein as demonstrated by immunoprecipitation of 125I-surface-labeled cells. Although MAbII-1 reacted with detergent-solubilized protein, it did not react with sodium-dodecyl sulfate-denatured protein by immunoblot assay. This protein was not exposed on the surface of phase I cells, but chloroform-methanol extraction of phase I cells exposed the phase II epitope.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Baca O. G., Paretsky D. Q fever and Coxiella burnetii: a model for host-parasite interactions. Microbiol Rev. 1983 Jun;47(2):127–149. doi: 10.1128/mr.47.2.127-149.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Batteiger B., Newhall W. J., 5th, Jones R. B. The use of Tween 20 as a blocking agent in the immunological detection of proteins transferred to nitrocellulose membranes. J Immunol Methods. 1982 Dec 30;55(3):297–307. doi: 10.1016/0022-1759(82)90089-8. [DOI] [PubMed] [Google Scholar]
  3. Beher M., Pugsley A., Schnaitman C. Correlation between the expression of an Escherichia coli cell surface protein and the ability of the protein to bind to lipopolysaccharide. J Bacteriol. 1980 Jul;143(1):403–410. doi: 10.1128/jb.143.1.403-410.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Burton P. R., Kordová N., Paretsky D. Electron microscopic studies of the rickettsia Coxiella burneti: entry, lysosomal response, and fate of rickettsial DNA in L-cells. Can J Microbiol. 1971 Feb;17(2):143–150. doi: 10.1139/m71-025. [DOI] [PubMed] [Google Scholar]
  5. Ciampor F., Schramek S., Brezina R. Electron microscopy of ruthenium red-stained phase I and II Coxiella burneti. Acta Virol. 1972 Nov;16(6):503–506. [PubMed] [Google Scholar]
  6. Crăcea E., Dumitrescu S., Novac S., Mihancea N. The influence of phase variation on the protective activity of C. burneti vaccines. Arch Roum Pathol Exp Microbiol. 1966 Dec;25(4):943–950. [PubMed] [Google Scholar]
  7. Ebeling W., Hennrich N., Klockow M., Metz H., Orth H. D., Lang H. Proteinase K from Tritirachium album Limber. Eur J Biochem. 1974 Aug 15;47(1):91–97. doi: 10.1111/j.1432-1033.1974.tb03671.x. [DOI] [PubMed] [Google Scholar]
  8. FISET P. Phase variation of Rickettsia (Coxiella) burneti; study of the antibody response in guinea pigs and rabbits. Can J Microbiol. 1957 Apr;3(3):435–445. doi: 10.1139/m57-046. [DOI] [PubMed] [Google Scholar]
  9. Fiset P., Ormsbee R. A., Silberman R., Peacock M., Spielman S. H. A microagglutination technique for detection and measurement of rickettsial antibodies. Acta Virol. 1969 Jan;13(1):60–66. [PubMed] [Google Scholar]
  10. Fiset P., Ormsbee R. A. The antibody response to antigens of Coxiella burneti. Zentralbl Bakteriol Orig. 1968 Apr;206(3):321–329. [PubMed] [Google Scholar]
  11. Fiset P., Wike D. A., Pickens E. G., Ormsbee R. A. An antigenic comparison of strains of Coxiella burneti. Acta Virol. 1971 Mar;15(2):161–166. [PubMed] [Google Scholar]
  12. Goding J. W. Use of staphylococcal protein A as an immunological reagent. J Immunol Methods. 1978;20:241–253. doi: 10.1016/0022-1759(78)90259-4. [DOI] [PubMed] [Google Scholar]
  13. Grey H. M., Hirst J. W., Cohn M. A new mouse immunoglobulin: IgG3. J Exp Med. 1971 Feb 1;133(2):289–304. doi: 10.1084/jem.133.2.289. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hackstadt T., Williams J. C. Biochemical stratagem for obligate parasitism of eukaryotic cells by Coxiella burnetii. Proc Natl Acad Sci U S A. 1981 May;78(5):3240–3244. doi: 10.1073/pnas.78.5.3240. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Halevy M., Goldwasser R. A. Differentiation of phase I and phase II Coxiella burneti by means of fluorescent antibody techniques. Isr J Med Sci. 1972 May;81(5):583–587. [PubMed] [Google Scholar]
  16. Kawakami M., Ihara I., Suzuki A., Harada Y. Properties of a new complement-dependent bactericidal factor specific for Ra chemotype salmonella in sera of conventional and germ-free mice. J Immunol. 1982 Nov;129(5):2198–2201. [PubMed] [Google Scholar]
  17. Krauss H., Schiefer H. G., Schmatz H. D. Ultrastructural investigations on surface structures involved in Coxiella burnetii phase variation. Infect Immun. 1977 Mar;15(3):890–896. doi: 10.1128/iai.15.3.890-896.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Köhler G., Milstein C. Continuous cultures of fused cells secreting antibody of predefined specificity. Nature. 1975 Aug 7;256(5517):495–497. doi: 10.1038/256495a0. [DOI] [PubMed] [Google Scholar]
  19. Köhler G., Milstein C. Derivation of specific antibody-producing tissue culture and tumor lines by cell fusion. Eur J Immunol. 1976 Jul;6(7):511–519. doi: 10.1002/eji.1830060713. [DOI] [PubMed] [Google Scholar]
  20. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  21. ORMSBEE R. A., BELL E. J., LACKMAN D. B., TALLENT G. THE INFLUENCE OF PHASE ON THE PROTECTIVE POTENCY OF Q FEVER VACCINE. J Immunol. 1964 Mar;92:404–412. [PubMed] [Google Scholar]
  22. Osborn M. J., Wu H. C. Proteins of the outer membrane of gram-negative bacteria. Annu Rev Microbiol. 1980;34:369–422. doi: 10.1146/annurev.mi.34.100180.002101. [DOI] [PubMed] [Google Scholar]
  23. Perlmutter R. M., Hansburg D., Briles D. E., Nicolotti R. A., Davie J. M. Subclass restriction of murine anti-carbohydrate antibodies. J Immunol. 1978 Aug;121(2):566–572. [PubMed] [Google Scholar]
  24. Philip R. N., Casper E. A., Burgdorfer W., Gerloff R. K., Hughes L. E., Bell E. J. Serologic typing of rickettsiae of the spotted fever group by microimmunofluorescence. J Immunol. 1978 Nov;121(5):1961–1968. [PubMed] [Google Scholar]
  25. STOKER M. G., FISET P. Phase variation of the Nine Mile and other strains of Rickettsia burneti. Can J Microbiol. 1956 May;2(3):310–321. doi: 10.1139/m56-036. [DOI] [PubMed] [Google Scholar]
  26. STOKER M. G. Variation in complement-fixing activity of Rickettsia burneti during egg adaptation. J Hyg (Lond) 1953 Sep;51(3):311–321. doi: 10.1017/s0022172400015746. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Schramek S., Mayer H. Different sugar compositions of lipopolysaccharides isolated from phase I and pure phase II cells of Coxiella burnetii. Infect Immun. 1982 Oct;38(1):53–57. doi: 10.1128/iai.38.1.53-57.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. TIGERTT W. D., BENENSON A. S., GOCHENOUR W. S. Airborne Q fever. Bacteriol Rev. 1961 Sep;25:285–293. doi: 10.1128/br.25.3.285-293.1961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Tsai C. M., Frasch C. E. A sensitive silver stain for detecting lipopolysaccharides in polyacrylamide gels. Anal Biochem. 1982 Jan 1;119(1):115–119. doi: 10.1016/0003-2697(82)90673-x. [DOI] [PubMed] [Google Scholar]
  31. Williams J. C., Cantrell J. L. Biological and immunological properties of Coxiella burnetii vaccines in C57BL/10ScN endotoxin-nonresponder mice. Infect Immun. 1982 Mar;35(3):1091–1102. doi: 10.1128/iai.35.3.1091-1102.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Williams J. C., Peacock M. G., McCaul T. F. Immunological and biological characterization of Coxiella burnetii, phases I and II, separated from host components. Infect Immun. 1981 May;32(2):840–851. doi: 10.1128/iai.32.2.840-851.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]

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