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. 1984 Feb;19(2):230–234. doi: 10.1128/jcm.19.2.230-234.1984

Enzyme-linked immunosorbent assay with a monoclonal antibody for detecting group A meningococcal antigens in cerebrospinal fluid.

R J Sugasawara, C M Prato, J E Sippel
PMCID: PMC271027  PMID: 6142055

Abstract

Hybridomas were produced from spleen cells of BALB/c mice immunized with a membrane preparation from Neisseria meningitidis group A strain 4402 and S194/5.XXOBU.14 myeloma cells. The hybridomas were screened for secretion of antibodies suitable for an enzyme-linked immunosorbent assay (ELISA) diagnostic for group A meningococcal meningitis. One hybridoma antibody, 3G7, was directed against the pilus protein. This antibody bound to all six lipopolysaccharide and protein group A meningococcal serotyping strains, as well as to meningococcal strains from serogroups C, W135, and Y, but not to a strain of Escherichia coli, Haemophilus influenzae type b, or to two or more strains of Streptococcus pneumoniae, Neisseria gonorrhoeae, and Salmonella typhi. The ELISA used on antibody, antigen, antibody-conjugate sandwich. Rabbit anti-meningococcal serum was the coating antibody for the antibody sandwich, cerebrospinal fluids contained the bacterial antigens, and 3G7-alkaline phosphatase conjugate was the detecting antibody. The monoclonal antibody conjugate ELISA system was able to detect group A meningococcal antigens in 21 of 25 cerebrospinal fluid specimens that were positive in an immune rabbit serum conjugate ELISA; cerebrospinal fluid samples from patients with Haemophilus meningitis served as the controls. Counterimmunoelectrophoresis detected meningococcal antigens in 16 of the same 25 cerebrospinal fluid samples.

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Selected References

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

  1. Brinton C. C., Jr The structure, function, synthesis and genetic control of bacterial pili and a molecular model for DNA and RNA transport in gram negative bacteria. Trans N Y Acad Sci. 1965 Jun;27(8):1003–1054. doi: 10.1111/j.2164-0947.1965.tb02342.x. [DOI] [PubMed] [Google Scholar]
  2. Buchanan T. M. Antigenic heterogeneity of gonococcal pili. J Exp Med. 1975 Jun 1;141(6):1470–1475. doi: 10.1084/jem.141.6.1470. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Burnette W. N. "Western blotting": electrophoretic transfer of proteins from sodium dodecyl sulfate--polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal Biochem. 1981 Apr;112(2):195–203. doi: 10.1016/0003-2697(81)90281-5. [DOI] [PubMed] [Google Scholar]
  4. Coller H. A., Coller B. S. Statistical analysis of repetitive subcloning by the limiting dilution technique with a view toward ensuring hybridoma monoclonality. Hybridoma. 1983;2(1):91–96. doi: 10.1089/hyb.1983.2.91. [DOI] [PubMed] [Google Scholar]
  5. Craven D. E., Peppler M. S., Frasch C. E., Mocca L. F., McGrath P. P., Washington G. Adherence of isolates of Neisseria meningitidis from patients and carriers to human buccal epithelial cells. J Infect Dis. 1980 Oct;142(4):556–568. doi: 10.1093/infdis/142.4.556. [DOI] [PubMed] [Google Scholar]
  6. Edwards E. A. Immunologic investigations of meningococcal disease. I. Group-specific Neisseria meningitidis antigens present in the serum of patients with fulminant meningococcemia. J Immunol. 1971 Feb;106(2):314–317. [PubMed] [Google Scholar]
  7. Engvall E., Perlmann P. Enzyme-linked immunosorbent assay (ELISA). Quantitative assay of immunoglobulin G. Immunochemistry. 1971 Sep;8(9):871–874. doi: 10.1016/0019-2791(71)90454-x. [DOI] [PubMed] [Google Scholar]
  8. Frasch C. E., Gotschlich E. C. An outer membrane protein of Neisseria meningitidis group B responsible for serotype specificity. J Exp Med. 1974 Jul 1;140(1):87–104. doi: 10.1084/jem.140.1.87. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hermodson M. A., Chen K. C., Buchanan T. M. Neisseria pili proteins: amino-terminal amino acid sequences and identification of an unusual amino acid. Biochemistry. 1978 Feb 7;17(3):442–445. doi: 10.1021/bi00596a010. [DOI] [PubMed] [Google Scholar]
  10. Higashi G. I., Sippel J. E., Girgis N. I., Hassan A. Counterimmunoelectrophoresis: an adjunct to bacterial culture in the diagnosis of meningococcal meningitis. Scand J Infect Dis. 1974;6(3):233–235. doi: 10.3109/inf.1974.6.issue-3.04. [DOI] [PubMed] [Google Scholar]
  11. Kennett R. H. Cell fusion. Methods Enzymol. 1979;58:345–359. doi: 10.1016/s0076-6879(79)58149-x. [DOI] [PubMed] [Google Scholar]
  12. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  13. 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]
  14. Leinonen M., Herva E. The latex agglutination test for the diagnosis of meningococcal and haemophilus influenzae meningitis. Scand J Infect Dis. 1977;9(3):187–191. doi: 10.3109/inf.1977.9.issue-3.06. [DOI] [PubMed] [Google Scholar]
  15. Olcén P. Serological methods for rapid diagnosis of haemophilus influenzae, Neisseria meningitidis and Streptococcus pneumoniae in cerebrospinal fluid: a comparison of Co-agglutination, immunofluorescence and immunoelectroosmophoresis. Scand J Infect Dis. 1978;10(4):283–289. doi: 10.3109/inf.1978.10.issue-4.05. [DOI] [PubMed] [Google Scholar]
  16. Sippel J. E., Girgis N. I. Meningococcal infection in Egypt: laboratory findings in meningitis patients and the prevalence of pharyngeal infection in patients and contacts. Am J Trop Med Hyg. 1978 Sep;27(5):980–985. doi: 10.4269/ajtmh.1978.27.980. [DOI] [PubMed] [Google Scholar]
  17. Sippel J. E., Quan A. Homogeneity of protein serotype antigens in Neisseria meningitidis group A. Infect Immun. 1977 May;16(2):623–627. doi: 10.1128/iai.16.2.623-627.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Sippel J. E., Voller A. Detection of Neisseria meningitidis cell envelope antigen by enzyme-linked immunosorbent assay in patients with meningococcal disease. Trans R Soc Trop Med Hyg. 1980;74(5):644–648. doi: 10.1016/0035-9203(80)90156-x. [DOI] [PubMed] [Google Scholar]
  19. 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]
  20. Whittle H. C., Tugwell P., Egler L. J., Greenwood B. M. Rapid bacteriological diagnosis of pyogenic meningitis by latex agglutination. Lancet. 1974 Sep 14;2(7881):619–621. doi: 10.1016/s0140-6736(74)91943-6. [DOI] [PubMed] [Google Scholar]
  21. Yolken R. H. Enzyme immunoassays for the detection of infectious antigens in body fluids: current limitations and future prospects. Rev Infect Dis. 1982 Jan-Feb;4(1):35–68. doi: 10.1093/clinids/4.1.35. [DOI] [PubMed] [Google Scholar]
  22. Zollinger W. D., Mandrell R. E. Type-specific antigens of group A Neisseria meningitidis: lipopolysaccharide and heat-modifiable outer membrane proteins. Infect Immun. 1980 May;28(2):451–458. doi: 10.1128/iai.28.2.451-458.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. de StGroth S. F., Scheidegger D. Production of monoclonal antibodies: strategy and tactics. J Immunol Methods. 1980;35(1-2):1–21. doi: 10.1016/0022-1759(80)90146-5. [DOI] [PubMed] [Google Scholar]

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