Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1982 Mar;79(5):1673–1677. doi: 10.1073/pnas.79.5.1673

Isolation and characterization of a monoclonal antibody against the saxitoxin-binding component from the electric organ of the eel Electrophorus electricus.

H P Moore, L C Fritz, M A Raftery, J P Brockes
PMCID: PMC346038  PMID: 6280195

Abstract

A monoclonal hybridoma cell line secreting antibody against the saxitoxin-binding component from the eel Electrophorus electricus has been isolated. The specificity of this monoclonal antibody was established by (i) its ability to immunoprecipitate bound [3H]saxitoxin from a detergent extract of electroplax membranes in a dose-dependent manner, (ii) the inability of unrelated monoclonal antibodies to immunoprecipitate the toxin-binding activity in a similar assay, and (iii) the ability of excess unlabeled tetrodotoxin to displace [3H]saxitoxin from the immunoprecipitated component. The antibody is of the subclass IgG1 and binds specifically to a polypeptide component of Mr approximately 250,000 on NaDodSO4/polyacrylamide gels. The antigenic determinant is associated with the same polypeptide component throughout the purification procedure, indicating that this component is not a result of artifactual aggregation or degradation during isolation. We conclude that the 250,000-dalton polypeptide is part of the saxitoxin binding/sodium channel protein in the native electroplax membrane.

Full text

PDF
1673

Images in this article

1676Image
on p.1676

Selected References

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

  1. Agnew W. S., Levinson S. R., Brabson J. S., Raftery M. A. Purification of the tetrodotoxin-binding component associated with the voltage-sensitive sodium channel from Electrophorus electricus electroplax membranes. Proc Natl Acad Sci U S A. 1978 Jun;75(6):2606–2610. doi: 10.1073/pnas.75.6.2606. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Agnew W. S., Moore A. C., Levinson S. R., Raftery M. A. Identification of a large molecular weight peptide associated with a tetrodotoxin binding protein from the electroplax of Electrophorus electricus. Biochem Biophys Res Commun. 1980 Feb 12;92(3):860–866. doi: 10.1016/0006-291x(80)90782-2. [DOI] [PubMed] [Google Scholar]
  3. Barchi R. L., Cohen S. A., Murphy L. E. Purification from rat sarcolemma of the saxitoxin-binding component of the excitable membrane sodium channel. Proc Natl Acad Sci U S A. 1980 Mar;77(3):1306–1310. doi: 10.1073/pnas.77.3.1306. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Beneski D. A., Catterall W. A. Covalent labeling of protein components of the sodium channel with a photoactivable derivative of scorpion toxin. Proc Natl Acad Sci U S A. 1980 Jan;77(1):639–643. doi: 10.1073/pnas.77.1.639. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Brockes J. P., Fryxell K. J., Lemke G. E. Studies on cultured Schwann cells: the induction of myelin synthesis, and the control of their proliferation by a new growth factor. J Exp Biol. 1981 Dec;95:215–230. doi: 10.1242/jeb.95.1.215. [DOI] [PubMed] [Google Scholar]
  6. Burridge K. Direct identification of specific glycoproteins and antigens in sodium dodecyl sulfate gels. Methods Enzymol. 1978;50:54–64. doi: 10.1016/0076-6879(78)50007-4. [DOI] [PubMed] [Google Scholar]
  7. Catterall W. A. Neurotoxins that act on voltage-sensitive sodium channels in excitable membranes. Annu Rev Pharmacol Toxicol. 1980;20:15–43. doi: 10.1146/annurev.pa.20.040180.000311. [DOI] [PubMed] [Google Scholar]
  8. Engvall E., Perlmann P. Enzyme-linked immunosorbent assay, Elisa. 3. Quantitation of specific antibodies by enzyme-labeled anti-immunoglobulin in antigen-coated tubes. J Immunol. 1972 Jul;109(1):129–135. [PubMed] [Google Scholar]
  9. Goldin S. M., Rhoden V., Hess E. J. Molecular characterization, reconstitution, and "transport-specific fractionation" of the saxitoxin binding protein/Na+ gate of mammalian brain. Proc Natl Acad Sci U S A. 1980 Nov;77(11):6884–6888. doi: 10.1073/pnas.77.11.6884. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hartshorne R. P., Catterall W. A. Purification of the saxitoxin receptor of the sodium channel from rat brain. Proc Natl Acad Sci U S A. 1981 Jul;78(7):4620–4624. doi: 10.1073/pnas.78.7.4620. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. 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]
  12. 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]
  13. Levinson S. R., Curatalo C. J., Reed J., Raftery M. A. A rapid and precise assay for tetrodotoxin binding to detergent extracts of excitable tissues. Anal Biochem. 1979 Oct 15;99(1):72–84. doi: 10.1016/0003-2697(79)90045-9. [DOI] [PubMed] [Google Scholar]
  14. Ritchie J. M., Rogart R. B., Strichartz G. R. A new method for labelling saxitoxin and its binding to non-myelinated fibres of the rabbit vagus, lobster walking leg, and garfish olfactory nerves. J Physiol. 1976 Oct;261(2):477–494. doi: 10.1113/jphysiol.1976.sp011569. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES