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. 1975 Dec;72(12):4759–4763. doi: 10.1073/pnas.72.12.4759

Isolation of antibodies specific to sickle hemoglobin by affinity chromatography using a synthetic peptide.

N S Young, J G Curd, A Eastlake, B Furie, A N Schechter
PMCID: PMC388810  PMID: 813215

Abstract

Antibodies to hemoglobin have been studied with a radioimmunoassay which employs [14C]carbamylated (= carbamolyated) hemoglobin S. An antiserum raised against hemoglobin S, which initially discriminated poorly between hemoglobins S and A, was fractionated by absorption to a column of Sepharose to which a synthetic peptide corresponding to the first 13 amino-acid residues of the beta chain of sickle hemoglobin had been covalently bound. A subpopulation of the antiserum was eluted from this column with 4 M guanidine - HC1. These antibodies showed binding to hemoglobin S but not to hemoglobin A and this interaction could be inhibited by the synthetic peptide. These antibodies, of demonstrated fine structural specificity, may be useful in the detection of sickle hemoglobin and in the study of its structure in solution.

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Selected References

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

  1. Arnon R., Maron E., Sela M., Anfinsen C. B. Antibodies reactive with native lysozyme elicited by a completely synthetic antigen. Proc Natl Acad Sci U S A. 1971 Jul;68(7):1450–1455. doi: 10.1073/pnas.68.7.1450. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Corley L., Sachs D. H., Anfinsen C. B. Rapid solid-phase synthesis of bradykinin. Biochem Biophys Res Commun. 1972 Jun 28;47(6):1353–1359. doi: 10.1016/0006-291x(72)90221-5. [DOI] [PubMed] [Google Scholar]
  3. Cuatrecasas P., Wilchek M., Anfinsen C. B. Selective enzyme purification by affinity chromatography. Proc Natl Acad Sci U S A. 1968 Oct;61(2):636–643. doi: 10.1073/pnas.61.2.636. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Fearney F. J., Leung C. Y., Young J. D., Benjamini E. The specificity of antibodies to a peptide determinant of the tobacco mosaic virus protein induced by immunization with the peptide conjugate. Biochim Biophys Acta. 1971 Sep 28;243(3):509–514. doi: 10.1016/0005-2795(71)90024-9. [DOI] [PubMed] [Google Scholar]
  5. Furie B., Schechter A. N., Sachs D. H., Anfinsen C. B. Antibodies to the unfolded form of a helix-rich region in staphylococcal nuclease. Biochemistry. 1974 Apr 9;13(8):1561–1566. doi: 10.1021/bi00705a001. [DOI] [PubMed] [Google Scholar]
  6. Huisman T. H., Dozy A. M. Studies on the heterogeneity of hemoglobin. IX. The use of Tris(hydroxymethyl)aminomethanehcl buffers in the anion-exchange chromatography of hemoglobins. J Chromatogr. 1965 Jul;19(1):160–169. doi: 10.1016/s0021-9673(01)99434-8. [DOI] [PubMed] [Google Scholar]
  7. Kilmartin J. V., Rossi-Bernardi L. The binding of carbon dioxide by horse haemoglobin. Biochem J. 1971 Aug;124(1):31–45. doi: 10.1042/bj1240031. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. LAPRESLE C., WEBB T. ISOLATION AND STUDY OF A FRAGMENT OF HUMAN SERUM ALBUMIN CONTAINING ONE OF THE ANTIGENIC SITES OF THE WHOLE MOLECULE. Biochem J. 1965 Apr;95:245–251. doi: 10.1042/bj0950245. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Maron E., Shiozawa C., Arnon R., Sela M. Chemical and immunological characterization of a unique antigenic region in lysozyme. Biochemistry. 1971 Mar 2;10(5):763–771. doi: 10.1021/bi00781a007. [DOI] [PubMed] [Google Scholar]
  10. Nisonoff A., Reichlin M., Margoliash E. Immunological activity of cytochrome c. II. Localization of a major antigenic determinant of human cytochrome c. J Biol Chem. 1970 Mar 10;245(5):940–946. [PubMed] [Google Scholar]
  11. Noble R. W., Reichlin M., Schreiber R. D. Studies on antibodies directed toward single antigenic sites on globular proteins. Biochemistry. 1972 Aug 29;11(18):3326–3332. doi: 10.1021/bi00768a004. [DOI] [PubMed] [Google Scholar]
  12. Perutz M. F. Stereochemistry of cooperative effects in haemoglobin. Nature. 1970 Nov 21;228(5273):726–739. doi: 10.1038/228726a0. [DOI] [PubMed] [Google Scholar]
  13. REICHLIN M., BUCCI E., ANTONINI E., WYMAN J., ROSSI-FANELLI A. THE IMMUNOCHEMICAL DIFFERENCE BETWEEN HORSE OXY- AND DEOXYHAEMOGLOBIN. J Mol Biol. 1964 Sep;9:785–788. doi: 10.1016/s0022-2836(64)80184-4. [DOI] [PubMed] [Google Scholar]
  14. Reichlin M. Localizing antigenic determinants in human haemoglobin with mutants: molecular correlations of immunological tolerance. J Mol Biol. 1972 Mar 14;64(2):485–496. doi: 10.1016/0022-2836(72)90512-8. [DOI] [PubMed] [Google Scholar]
  15. Rowley P. T., Doherty R. A., Rosecrans C., Cernichiari E. Sickle hemoglobin: a specific radioimmunoassay. Blood. 1974 Apr;43(4):607–611. [PubMed] [Google Scholar]
  16. Sachs D. H., Schechter A. N., Eastlake A., Anfinsen C. B. An immunologic approach to the conformational equilibria of polypeptides. Proc Natl Acad Sci U S A. 1972 Dec;69(12):3790–3794. doi: 10.1073/pnas.69.12.3790. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Sachs D. H., Schechter A. N., Eastlake A., Anfinsen C. B. Antibodies to a distinct antigenic determinant of staphylococcal nuclease. J Immunol. 1972 Dec;109(6):1300–1310. [PubMed] [Google Scholar]
  18. Schreiber R. D., Noble R. W., Reichlin M. Restriction of heterogeneity of goat antibodies specific for human hemoglobin SS. J Immunol. 1975 Jan;114(1 Pt 1):170–175. [PubMed] [Google Scholar]
  19. Ungar-Waron H., Hurwitz E., Jaton J. C., Sela M. Antibodies elicited with conjugates of nucleosides with synthetic polypeptides. Biochim Biophys Acta. 1967 May 30;138(3):513–531. doi: 10.1016/0005-2787(67)90548-5. [DOI] [PubMed] [Google Scholar]
  20. WASSERMAN E., LEVINE L. Quantitative micro-complement fixation and its use in the study of antigenic structure by specific antigen-antibody inhibition. J Immunol. 1961 Sep;87:290–295. [PubMed] [Google Scholar]

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