Combining Site Specificity and Idiotypy

A Study of Antidigoxin and Antiarsonate Antibodies
  • Edgar Haber
  • Michael N. Margolies


We are now rather close to understanding the mechanisms whereby antibody combining site diversity is generated. As this story has unfolded over the past several years, a number of unique genetic mechanisms have been uncovered and many of the issues that fueled controversy among laboratories have been resolved.


Light Chain Heavy Chain Somatic Mutation Antigen Binding Partial Amino Acid Sequence 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    Brack, C., Hirama, M., Lenhard-Schuller, R., and Tonegawa, S., 1978, A complete immunoglobulin gene is created by somatic recombination, Cell 15:1–14.PubMedCrossRefGoogle Scholar
  2. 2.
    Sakano, H., Kurosawa, Y., Weigert, M., and Tonegawa, S., 1981, Identification and nucleotide sequence of a diversity DNA segment (D) of immunoglobulin heavy-chain genes, Nature 290:562–565.PubMedCrossRefGoogle Scholar
  3. 3.
    Seidman, J. G., Leder, A., Nau, M., Norman, B., and Leder, P., 1978, Antibody diversity: The structure of cloned immunoglobulin genes suggests a mechanism for generating new sequences, Science 202:11–17.PubMedCrossRefGoogle Scholar
  4. 4.
    Seidman, J. G., Max, E. E., and Leder, P., 1979, A kappa-immunoglobulin gene is formed by site-specific recombination without further somatic mutation, Nature 280:370–375.PubMedCrossRefGoogle Scholar
  5. 5.
    Early, P., Huang, H., Davis, M., Calame, K., and Hood, L., 1980, An immunoglobulin heavy chain variable region gene is generated from three segments of DNA: VH, D, and JH, Cell 19:981–992.PubMedCrossRefGoogle Scholar
  6. 6.
    Weigert, M., Gatmaitan, L., Loh, E., Schilling, J., and Hood, L., 1978, Rearrangement of genetic information may produce immunoglobulin diversity, Nature 276:785–790.PubMedCrossRefGoogle Scholar
  7. 7.
    Tonegawa, S., 1983, Somatic generation of antibody diversity, Nature 302:575–581.PubMedCrossRefGoogle Scholar
  8. 8.
    Gearhart, P. J., 1983, Effect of somatic mutation on antibody affinity, Ann. N.Y. Acad. Sci. 418:171–176.PubMedCrossRefGoogle Scholar
  9. 9.
    Clarke, S. H., Claflin, J. L., and Rudikoff, S., 1982, Polymorphisms in immunoglobulin heavy chains suggesting gene conversion,Proc. Natl. Acad. Sci. USA 79:3280–3284.PubMedCrossRefGoogle Scholar
  10. 10.
    Hochman, J., Inbar, D., and Givol, D., 1976, An active antibody fragment (Fv) composed of the variable portions of heavy and light chains, Biochemistry 12:1130–1135.CrossRefGoogle Scholar
  11. 11.
    Marquart, M., Deisenhofer, J., Huber, R., and Palm, W., 1980, Crystallographic refinement and atomic models of the intact immunoglobulin molecule Kol and its antigen-binding fragment at 3.0 A and 1.0 A resolution, J. Mol. Biol. 141:369–391.PubMedCrossRefGoogle Scholar
  12. 12.
    Segal, D. M., Padlan, E. A., Cohen, G. H., Rudikoff, S., Potter, M., and Davies, D. R., 1974, The three- dimensional structure of phosphorylcholine-binding mouse immunoglobulin Fab and the nature of the antigen binding site, Proc. Natl. Acad. Sci. USA 71:4298–4302.PubMedCrossRefGoogle Scholar
  13. 13.
    Saul, F. A., Amzel, L. M., and Poljak, R. J., 1978, Preliminary refinement and structural analysis of the Fab fragment from human immunoglobulin New at 2.0 A resolution, J. Biol. Chem. 253:585–595.PubMedGoogle Scholar
  14. 14.
    Novotny, J., Bruccoleri, R., Newell, J., Murphy, D., Haber, E., and Karplus, M., 1983, Molecular anatomy of the antibody combining site, J. Biol. Chem. 258:14433–14437.PubMedGoogle Scholar
  15. 15.
    Kabat, E. A., 1966, The nature of an antigenic determinant, J. Immunol. 97:1–11.PubMedGoogle Scholar
  16. 16.
    Go, K., Kartha, G., and Chen, J. P., 1980, Structure of digoxin, Acta Crystallogr. Sect. B 36:1811–1819.CrossRefGoogle Scholar
  17. 17.
    Fieser, L. F., and Fieser, M., 1959,Steroids, Reinhold, New York, p. 727.Google Scholar
  18. 18.
    Smith, T. W., Lloyd, B. L., Spicer, N., and Haber, E., 1979, Immunogenicity and kinetics of distribution and elimination of sheep digoxin-specific IgG and Fab fragments in the rabbit and baboon, Clin. Exp. Immunol. 36:384–396.PubMedGoogle Scholar
  19. 19.
    Curd, J., Smith, T. W., Jaton, J. C., and Haber, E., 1971, The isolation of digoxin-specific antibody and its use in reversing the effects of digoxin, Proc. Natl. Acad. Sci. USA 68:2401–2406.PubMedCrossRefGoogle Scholar
  20. 20.
    Smith, T. W., Butler, V. P., Jr., Haber, E., Fozzard, H., Marcus, F. I., Bremner, W. F., Schulman, I. C., and Phillips, A., 1982, Treatment of life-threatening digitalis intoxication with digoxin-specific Fab antibody fragments: Experience in 26 cases, N. Engl. J. Med. 307:1357–1362.PubMedCrossRefGoogle Scholar
  21. 21.
    Aeberhard, P., Butler, V. P., Smith, T. W., Haber, E., Tse Eng, D., Brau, J., Chalom, A., Glatt, B., Thebaut, J. F., Delangenhagen, B., and Morin, B., 1980, Le traitement d’une intoxication digitalique massive (20 mg de digitoxine) par les anticorps anti-digoxine fractionnes (Fab), Arch. Mai. Coeur Vaiss. 73:1471–1478.Google Scholar
  22. 22.
    Bismuth, C., Gaultier, M., Conso, F., and Efthymiou, M. L., 1973, Hyperkalemia in acute digitalis poisoning: Prognostic significance and therapeutic implications, Clin. Toxicol. 6:153–162.PubMedCrossRefGoogle Scholar
  23. 23.
    Margolies, M. N., Mudgett-Hunter, M., Smith, T. W., Novotny, J., and Haber, E., 1981, Monoclonal antibodies to the cardiac glycoside digoxin, in: Monoclonal Antibodies and T Cell Hybridomas (G. Ham- merling, U. Hammerling, and J. F. Kearney, eds.), Elsevier/North-Holland, Amsterdam, pp. 367–374.Google Scholar
  24. 24.
    Lechat, P., Mudgett-Hunter, M., Margolies, M. N., Haber, E., and Smith, T. W., 1984, Reversal of lethal digoxin toxicity in guinea pigs using monoclonal antibodies and Fab fragments, f. Phys. Exp. Ther. (in press).Google Scholar
  25. 25.
    Mudgett-Hunter, M., Margolies, M. N., Ju, A., and Haber, E., 1982, High-affinity monoclonal antibodies to the cardiac glycoside digoxin, J. Immunol. 129:1165–1172.Google Scholar
  26. 26.
    Ochs, H. R., and Smith, T. W., 1977, Reversal of advanced digitoxin toxicity and modification of pharmacokinetics by specific antibodies and Fab fragments,J. Clin. Invest. 60:1303–1313.PubMedCrossRefGoogle Scholar
  27. 27.
    Smith, T. W., Butler, V. P., and Haber, E., 1970, Characterization of antibodies of high affinity and specificity to the digitalis glycoside digoxin, Biochemistry 9:331–337.PubMedCrossRefGoogle Scholar
  28. 28.
    Sigal, N. H., Gearhart, P. J., and Klinman, N. R., 1975, The frequency of phosphorylcholine-specific B cells in conventional and germ free Balb/C mice, J. Immunol. 114:1354–1358.PubMedGoogle Scholar
  29. 29.
    Sigal, N., 1977, The frequency of/?-azophenylarsonate and dimethylamino-propthalene sulfonyl-specific B cells in neonatal and adult Balb/C mice, J. Immunol. 119:1129–1133.PubMedGoogle Scholar
  30. 30.
    Kohler, G., and Milstein, C., 1975, Continuous cultures of fused cells secreting antibody of predefined specificity, Nature 256:494–497.CrossRefGoogle Scholar
  31. 31.
    Shulman, M., Wilde, C. D., and Kohler, G., 1978, A better cell line for making hybridomas secreting specific antibodies, Nature 276:269–270.PubMedCrossRefGoogle Scholar
  32. 32.
    Klinman, N. R., Pickard, A. R., Sigal, N. H., Gearhart, P. J., Metcalf, E. S., and Pierce, S. K., 1976, Assessing B cell diversification by antigen receptor and precursor cell analysis,Ann. Immunol. (Paris) 127C:489–502.Google Scholar
  33. 33.
    Smith, T. W., 1972, Ouabain specific antibodies and immunochemical properties and reversal of Na-K- ATPase inhibition, J. Clin. Invest. 51:1583–1593.PubMedCrossRefGoogle Scholar
  34. 34.
    Brauer, A. W., Margolies, M. N., and Haber, E., 1975, The application of 0.1 M Quadrol to the micro- sequence of proteins and the sequence of tryptic peptides, Biochemistry 14:3029–3035.PubMedCrossRefGoogle Scholar
  35. 35.
    Novotny, J., and Margolies, M. N., 1983, Amino acid sequence of light chain variable region from a mouse anti-digoxin hybridoma antibody, Biochemistry 22:1153–1158.PubMedCrossRefGoogle Scholar
  36. 36.
    Margolies, M. N., and Brauer, A. W., 1978, Protein microsequencing using high pressure liquid chromatography of phenylthiohydantoin amino acids,J. Chromatogr. 148:429–439.CrossRefGoogle Scholar
  37. 37.
    Margolies, M. N., Brauer, A. W., Oman, C. L., Klapper, D. G., and Horn, M. J., 1982, Improved automatic conversion for use with a liquid-phase sequencer, in: Proceedings of IVth International Conference on Methods in Protein Sequence Analysis (M. Elzinga, ed.), Humana Press, Clifton, N.J., pp. 189–203.CrossRefGoogle Scholar
  38. 38.
    Clarke, S. H., Claflin, J. L., Potter, M., and Rudikoff, S., 1982, Polymorphisms in anti-phosphocholine antibodies reflecting evolution of immunoglobulin gene families, J. Exp. Med. 157:98–113.CrossRefGoogle Scholar
  39. 39.
    Rose, D. R., Seaton, B. A., Petsko, G. A., Novotny, J., Margolies, M. N., Locke, E., and Haber, E., 1983, Crystallization of the Fab fragment of a monoclonal anti-digoxin antibody and its complex with digoxin, J. Mol. Biol. 164:203–206.CrossRefGoogle Scholar
  40. 40.
    Kuettner, M. G., Wang, A. L., and Nisonoff, A., 1972, Quantitative investigations of idiotypic antibodies. VI. Idiotypic specificity as a potential genetic marker for the variable regions of mouse immunoglobulin polypeptide chains, J. Exp. Med. 135:579–595.PubMedCrossRefGoogle Scholar
  41. 41.
    Pawlak, L. L., Hart, D. A., and Nisonoff, A., 1973, Requirements for prolonged suppression of an idiotypic specificity in adult mice, J. Exp. Med. 137:1442–1458.PubMedCrossRefGoogle Scholar
  42. 42.
    Jerne, N. K., 1974, Towards a network theory of the immune system, Ann. Immunol. (Inst. Pasteur) 125C:373–389.Google Scholar
  43. 43.
    Capra, J. D., and Nisonoff, A., 1979, Structural studies on induced antibodies with defined idiotypic specificities. VII. The complete amino acid sequence of the heavy chain variable region of anti-jfr-azophenylar- sonate antibodies from A/J mice bearing a cross-reactive idiotype, J. Immunol. 123:279–284.PubMedGoogle Scholar
  44. 44.
    Marshak-Rothstein, A., Siekevitz, M., Margolies, M. N., Mudgett-Hunter, M., and Gefter, M. L., 1980, Hybridoma proteins expressing the predominant idiotype of the antiphenylarsonate response of the A/J mouse, Proc. Natl. Acad. Sci. USA 77:1120–1124.PubMedCrossRefGoogle Scholar
  45. 45.
    Marshak-Rothstein, A., Margolies, M. N., Riblet, R., and Gefter, M. L., 1981, Specificity of idiotype suppression in the A/J anti-azophenylarsonate system, in: Immunoglobulin Idiotypes (C. Janeway, E. E. Sercarz, and H. Wigzell, eds.), Academic Press, New York, pp. 739–749.Google Scholar
  46. 46.
    Rothstein, T. L., Margolies, M. N., Gefter, M. L., and Marshak-Rothstein, A., 1983, Fine specificity of idiotype suppression in the A/J anti-azophenylarsonate response,J. Exp. Med. 157:795–800.PubMedCrossRefGoogle Scholar
  47. 47.
    Marshak-Rothstein, A., Benedetto, J. D., Kirsch, R. L., and Gefter, M. L., 1980, Unique determinants associated with hybridoma proteins expressing a cross-reactive idiotype: Frequency among individual immune sera, J. Immunol. 125:1987–1992.PubMedGoogle Scholar
  48. 48.
    Nelles, M. J., Gill-Pazaris, L. A., and Nisonoff, A., 1981, Monoclonal anti-idiotypic antibodies reactive with a highly conserved determinant on A/J serum anti-p-azophenylarsonate antibodies, J. Exp. Med. 154:1752–1763.PubMedCrossRefGoogle Scholar
  49. 49.
    Estess, P., Nisonoff, A., and Capra, J. D., 1979, Structural studies on induced antibodies with defined idiotypic specificities. VIII. NH2-terminal amino acid sequence analysis of the heavy and light chain variable regions of monoclonal anti-/?-azophenylarsonate antibodies from A/J mice differing with respect to a cross-reactive idiotype, Mol. Immunol. 16:1111–1116.PubMedCrossRefGoogle Scholar
  50. 50.
    Estess, P., Lamoyi, E., Nisonoff, A., and Capra, J. D., 1980, Structural studies on induced antibodies with defined idiotype specificities. IX. Framework differences in the heavy- and light-chain-variable regions of monoclonal anti-/?-azophenylarsonate antibodies from A/J mice differing with respect to a cross-reactive idiotype, J. Exp. Med. 151:863–875.PubMedCrossRefGoogle Scholar
  51. 51.
    Margolies, M. N., Marshak-Rothstein, A., and Gefter, M. L., 1981, Structural diversity among anti-p- azophenylarsonate monoclonal antibodies from A/J mice: Comparison of Id- and Id+ sequences, Mol. Immunol. 18:1065–1077.PubMedCrossRefGoogle Scholar
  52. 52.
    Siegelman, M., and Capra, J. D., 1981, Complete amino acid sequence of light chain variable regions derived from five monoclonal anti-jfr-azophenylarsonate antibodies differing with respect to a cross-reactive idiotype, Proc. Natl. Acad. Sci. USA 78:7679–7683.PubMedCrossRefGoogle Scholar
  53. 53.
    Ball, R. K., Chang, J. Y., Alkan, S. S., and Braun, D. G., 1983, The complete amino acid sequence of the light chain variable region of two monoclonal anti-jfr-azobenzene-arsonate antibodies bearing the cross- reactive idiotype, Mol. Immunol. 20:197–201.PubMedCrossRefGoogle Scholar
  54. 54.
    Siekevitz, M., Gefter, M. L., Brodeur, P., Riblet, R., and Marshak-Rothstein, A., 1982, The genetic basis of antibody production: The dominant anti-arsonate idiotype response of the strain A mouse, Eur. J. Immunol. 12:1023–1032.PubMedCrossRefGoogle Scholar
  55. 55.
    Siekevitz, M., Huang, S. Y., and Gefter, M. L., 1983, The genetic basis of antibody production: One variable region heavy chain gene encodes all molecules bearing the dominant anti-arsonate idiotype in the strain A mouse, Eur. J. Immunol. 13:123–132.PubMedCrossRefGoogle Scholar
  56. 56.
    Marshak-Rothstein, A., Margolies, M. N., Benedetto, J. D., and Gefter, M. L., 1981, Two structurally distinct and independently regulated families associated with the A/J response to azophenylarsonate, Eur. J. Immunol. 11:565–572.PubMedCrossRefGoogle Scholar
  57. 57.
    Milner, E. C. B., and Capra, J. D., 1982, VH families in the antibody response to jfr-azophenylarsonate: Correlation between serology and amino acid sequence,J. Immunol. 129:193–199.PubMedGoogle Scholar
  58. 58.
    Kurosawa, Y., and Tonegawa, S., 1982, Organization, structure and assembly of immunoglobulin heavy chain diversity DNA segments, J. Exp. Med. 155:201–218.PubMedCrossRefGoogle Scholar
  59. 59.
    Margolies, M. N., Juszczak, E. C., Near, R., Marshak-Rothstein, A., Rothstein, T. L., Sato, V. L., Siekevitz, M., Smith, J. A., Wysocki, L. J., and Gefter, M. L., 1983, Structural correlates of idiotypy in the arsonate system, Ann. N.Y. Acad. Sci. 418:48–64.PubMedCrossRefGoogle Scholar
  60. 60.
    Capra, J. D., Slaughter, C., Milner, E. C. B., Estess, P., and Tucker, P. W., 1982, The cross-reactive idiotype of A strain mice, serological and structural analysis, Immunol. Today 3:332–339.CrossRefGoogle Scholar
  61. 61.
    Wysocki, L. J., and Sato, V. L., 1981, The strain A anti-/?-azophenylarsonate major cross-reactive idiotypic family includes members with no reactivity towards -azophenylarsonate, Eur. J. Immunol. 11:832–839.PubMedCrossRefGoogle Scholar
  62. 62.
    Margolies, M. N., Wysocki, L. J., and Sato, V. L., 1983, Immunoglobulin idiotype and anti-anti-idiotype utilize the same variable region genes irrespective of antigen specificity, J. Immunol. 130:515–517.PubMedGoogle Scholar
  63. 63.
    Gefter, M. L., Margolies, M. N., Near, R., and Wysocki, L. J., 1984, Analysis of the anti-azo-benzene- arsonate response at the molecular level, Ann. Inst. Pasteur 135:17–30.Google Scholar
  64. 64.
    Milner, E. C. B., and Capra, J. D., 1983, Structural analysis of monoclonal anti-arsonate antibodies: Idiotypic specificities are determined by the heavy chain,Mol. Immunol. 20:39–46.PubMedCrossRefGoogle Scholar
  65. 65.
    Cannon, L. E., and Woodland, R. T., 1983, Rapid and sensitive procedure for assigning idiotype determinants to heavy or light chains: Application to idiotype associated with the major cross-reactive idiotype of A/J antiphenylarsonate antibody, Molec. Immunol. 20:1283–1288.CrossRefGoogle Scholar
  66. 66.
    Sakano, H., Maki, R., Kurosawa, Y., Roeder, W., and Tonegawa, S., 1980, Two types of somatic recombination necessary for the generation of complete immunogloublin heavy-chain genes, Nature 286:676–683.PubMedCrossRefGoogle Scholar
  67. 67.
    Cook, W. D., Rudikoff, S., Giusti, A., and Scharff, M. D., 1982, Somatic mutation in a cultured mouse myeloma cell affects antigen binding, Proc. Natl. Acad. Sci. USA 79:1240–1244.PubMedCrossRefGoogle Scholar
  68. 68.
    Rudikoff, S., Giusti, A. M., Cook, W. D., and Scharff, M. D., 1982, Single amino acid substitution altering antigen-binding specificity,Proc. Natl. Acad. Sci. USA 79:1979–1983.PubMedCrossRefGoogle Scholar
  69. 69.
    Near, R. I., Juszczak, E. C., Huang, S. Y., Sicari, S. A., Margolies, M. N., and Gefter, M. L., 1984, Expression and rearrangement of homologous immunoglobulin VH genes in two mouse strains, Proc. Natl. Acad. Sci. USA 81:2167–2171.PubMedCrossRefGoogle Scholar
  70. 70.
    Gill-Pazaris, L. A., Brown, A. R., and Nisonoff, A., 1979, The nature of idiotypes associated with anti-p- azophenylarsonate antibodies in A/J mice, Ann. Immunol. (Inst. Pasteur) 130C: 199–213.Google Scholar
  71. 71.
    Brown, A. R., and Nisonoff, A., 1981, An intrastrain cross-reactive idiotype associated with anti-jfr-azo- phenylarsonate antibodies of Balb/C mice, J. Immunol. 126:1263–1267.PubMedGoogle Scholar
  72. 72.
    Brown, A. R., Lamoyi, E., and Nisonoff, A., 1981, Relationship of idiotypes of the anti-/?-azophenylar- sonate antibodies of A/J and Balb/C mice, J. Immunol. 126:1268–1273.PubMedGoogle Scholar
  73. 73.
    Brown, A. R., 1983, Idiotypic heterogeneity of the cross-reactive idiotype associated with the anti-/?-azo- phenylarsonate antibodies of Balb/C mice,J. Immunol. 131:423–428.PubMedGoogle Scholar
  74. 74.
    Juszczak, E. C., Near, R., and Margolies, M. N., 1984, in preparation.Google Scholar
  75. 75.
    Juszczak, E. C., and Margolies, M. N., 1983, Amino acid sequence of the heavy chain variable region from the A/J mouse anti-arsonate monoclonal antibody 36–60 bearing a minor idiotype, Biochemistry 22:4291- 4296.PubMedCrossRefGoogle Scholar
  76. 76.
    Gearhart, P. J., Johnson, N. D., Douglas, R., and Hood, L., 1981, IgG antibodies to phosphorylcholine exhibit more diversity than their IgM counterparts, Nature 291:29–34.PubMedCrossRefGoogle Scholar
  77. 77.
    Rothstein, T. L., and Gefter, M. L., 1983, Affinity analysis of idiotype-positive and idiotype-negative arsonate-binding hybridoma proteins and Ars-immune sera,Mol. Immunol. 20:161–168.PubMedCrossRefGoogle Scholar
  78. 78.
    Lewis, G. K., Kaymakcalan, Z., Yao, J., and Goodman, J. W., 1983, Idiotype connectance between anti- arsonate and anti-dinitrophenyl responses in Balb/C mice, Ann. N.Y. Acad. Sci. 418:282–289.PubMedCrossRefGoogle Scholar
  79. 79.
    Frederick, W. A., and Baltimore, D., 1982, Joining of immunoglobulin heavy chain gene segments: Implications from a chromosome with evidence of 3 D-JH fusions, Proc. Natl. Acad. Sci. USA 79:4118–4122.CrossRefGoogle Scholar
  80. 80.
    Oi, V. T., Morrison, S. L., Herzenberg, L. A., and Berg, P., 1983, Immunoglobulin gene expression in transformed lymphoid cells, Proc. Natl. Acad. Sci. USA 89:825–829.CrossRefGoogle Scholar
  81. IUPAC-IUB Commission on Biochemical Nomenclature, 1968, J. Biol. Chem. 243:3557–3559.Google Scholar
  82. 82.
    Rabat, E. A., Wu, T. T., and Bilofsky, H., 1979, Sequences of immunoglobulin chains: Tabulation and analyses of amino acid sequences of precursors, V-regions, C-regions, J-chains and BP-microglobulins, NIH Publication 80–2008.Google Scholar
  83. 83.
    Siegelman, M., Slaughter, C., McCumber, L., Estess, P., and Capra, J. D., 1981, Primary structural studies of monoclonal A/J anti-arsonate antibodies differing with respect to a cross-reactive idiotype, in: Immunoglobulin Idiotypes (C. Janeway, E. E. Sercarz, and H. Wigzell, eds.), Academic Press, New York, pp. 135–158.Google Scholar
  84. 84.
    Sims, J., Rabbitts, T. H., Estess, P., Slaugher, C., Tucker, P. W., and Capra, J. D., 1982, Somatic mutation in genes for the variable portions of the immunoglobulin heavy chain, Science 216:309–310.PubMedCrossRefGoogle Scholar
  85. 85.
    Alkan, S. S., Knecht, R., and Braun, D. G., 1980, The cross-reactive idiotype of anti-4-azobenzene-arsonate hybridoma-derived antibodies in A/J mice constitutes multiple heavy chains, Z. Physiol. Chem. 361:191–195Google Scholar

Copyright information

© Plenum Press, New York 1984

Authors and Affiliations

  • Edgar Haber
    • 1
  • Michael N. Margolies
    • 2
  1. 1.Cardiac Unit, Department of MedicineMassachusetts General Hospital and Harvard Medical SchoolBostonUSA
  2. 2.Department of SurgeryMassachusetts General Hospital and Harvard Medical SchoolBostonUSA

Personalised recommendations