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. 1984 Aug;74(2):625–633. doi: 10.1172/JCI111460

Molecular properties of the Ro/SSA antigen and enzyme-linked immunosorbent assay for quantitation of antibody.

H Yamagata, J B Harley, M Reichlin
PMCID: PMC370515  PMID: 6746910

Abstract

Antibodies to the Ro/SSA antigen occur in patients with systemic lupus erythematosus and Sjögren's syndrome. An immunoaffinity method for the preparation of electrophoretically homogeneous Ro/SSA antigen is described. Several molecular properties of the antigen have been determined. The native RNA protein particle has a molecular weight of approximately 100,000 D determined by gel filtration. Sodium dodecyl sulfate-analysis of the purified Ro/SSA antigen and analysis by staining of bands with silver and Coomassie Blue, Western blotting, and RNAase treatment leads to a hypothesis for the structure of the particle in which an antigenic 60,000 protein is bound to 24,000-27,000 RNA molecules which are not antigenic. An enzyme-linked immunoabsorbent assay method for assay of anti-Ro/SSA is also described which sensitively measures antigen binding at dilutions of sera containing anti-Ro/SSA precipitins up to 10(7) fold. Normal sera on average have 10(3) less binding activity.

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Selected References

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  1. Acuto O., Hussey R. E., Fitzgerald K. A., Protentis J. P., Meuer S. C., Schlossman S. F., Reinherz E. L. The human T cell receptor: appearance in ontogeny and biochemical relationship of alpha and beta subunits on IL-2 dependent clones and T cell tumors. Cell. 1983 Oct;34(3):717–726. doi: 10.1016/0092-8674(83)90528-7. [DOI] [PubMed] [Google Scholar]
  2. Alspaugh M. A., Tan E. M. Antibodies to cellular antigens in Sjögren's syndrome. J Clin Invest. 1975 May;55(5):1067–1073. doi: 10.1172/JCI108007. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Alspaugh M., Maddison P. Resolution of the identity of certain antigen-antibody systems in systemic lupus erythematosus and Sjögren's syndrome: an interlaboratory collaboration. Arthritis Rheum. 1979 Jul;22(7):796–798. doi: 10.1002/art.1780220719. [DOI] [PubMed] [Google Scholar]
  4. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1016/0003-2697(76)90527-3. [DOI] [PubMed] [Google Scholar]
  5. Catalano M. A., Krick E. H., De Heer D. H., Nakamura R. M., Theofilopoulos A. N., Vaughan J. H. Metabolism of autologous and homologous IgG in rheumatoid arthritis. J Clin Invest. 1977 Aug;60(2):313–322. doi: 10.1172/JCI108779. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Clark G., Reichlin M., Tomasi T. B., Jr Characterization of a soluble cytoplasmic antigen reactive with sera from patients with systemic lupus erythmatosus. J Immunol. 1969 Jan;102(1):117–122. [PubMed] [Google Scholar]
  7. DAVIS B. J. DISC ELECTROPHORESIS. II. METHOD AND APPLICATION TO HUMAN SERUM PROTEINS. Ann N Y Acad Sci. 1964 Dec 28;121:404–427. doi: 10.1111/j.1749-6632.1964.tb14213.x. [DOI] [PubMed] [Google Scholar]
  8. England T. E., Bruce A. G., Uhlenbeck O. C. Specific labeling of 3' termini of RNA with T4 RNA ligase. Methods Enzymol. 1980;65(1):65–74. doi: 10.1016/s0076-6879(80)65011-3. [DOI] [PubMed] [Google Scholar]
  9. 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]
  10. Fessler L. I., Fessler J. H. Identification of the carboxyl peptides of mouse procollagen IV and its implications for the assembly and structure of basement membrane procollagen. J Biol Chem. 1982 Aug 25;257(16):9804–9810. [PubMed] [Google Scholar]
  11. Franco H. L., Weston W. L., Peebles C., Forstot S. L., Phanuphak P. Autoantibodies directed against sicca syndrome antigens in the neonatal lupus syndrome. J Am Acad Dermatol. 1981 Jan;4(1):67–72. doi: 10.1016/s0190-9622(81)70011-2. [DOI] [PubMed] [Google Scholar]
  12. Ingolia T. D., Craig E. A., McCarthy B. J. Sequence of three copies of the gene for the major Drosophila heat shock induced protein and their flanking regions. Cell. 1980 Oct;21(3):669–679. doi: 10.1016/0092-8674(80)90430-4. [DOI] [PubMed] [Google Scholar]
  13. Kato N., Hoshino H., Harada F. Nucleotide sequence of 4.5S RNA (C8 or hY5) from HeLa cells. Biochem Biophys Res Commun. 1982 Sep 16;108(1):363–370. doi: 10.1016/0006-291x(82)91875-7. [DOI] [PubMed] [Google Scholar]
  14. Kephart D. C., Hood A. F., Provost T. T. Neonatal lupus erythematosus: new serologic findings. J Invest Dermatol. 1981 Sep;77(3):331–333. doi: 10.1111/1523-1747.ep12482531. [DOI] [PubMed] [Google Scholar]
  15. 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]
  16. Lerner M. R., Boyle J. A., Hardin J. A., Steitz J. A. Two novel classes of small ribonucleoproteins detected by antibodies associated with lupus erythematosus. Science. 1981 Jan 23;211(4480):400–402. doi: 10.1126/science.6164096. [DOI] [PubMed] [Google Scholar]
  17. Maddison P. J., Provost T. T., Reichlin M. Serological findings in patients with "ANA-negative" systemic lupus erythematosus. Medicine (Baltimore) 1981 Mar;60(2):87–94. doi: 10.1097/00005792-198103000-00002. [DOI] [PubMed] [Google Scholar]
  18. Massague J., Guillette B. J., Czech M. P. Affinity labeling of multiplication stimulating activity receptors in membranes from rat and human tissues. J Biol Chem. 1981 Mar 10;256(5):2122–2125. [PubMed] [Google Scholar]
  19. Merril C. R., Goldman D., Sedman S. A., Ebert M. H. Ultrasensitive stain for proteins in polyacrylamide gels shows regional variation in cerebrospinal fluid proteins. Science. 1981 Mar 27;211(4489):1437–1438. doi: 10.1126/science.6162199. [DOI] [PubMed] [Google Scholar]
  20. Miyagawa S., Kitamura W., Yoshioka J., Sakamoto K. Placental transfer of anticytoplasmic antibodies in annular erythema of newborns. Arch Dermatol. 1981 Sep;117(9):569–572. [PubMed] [Google Scholar]
  21. Ohtsubo H., Ohmori H., Ohtsubo E. Nucleotide-sequence analysis of Tn3 (ap): implications for insertion and deletion. Cold Spring Harb Symp Quant Biol. 1979;43(Pt 2):1269–1277. doi: 10.1101/sqb.1979.043.01.144. [DOI] [PubMed] [Google Scholar]
  22. Poduslo J. F. Glycoprotein molecular-weight estimation using sodium dodecyl sulfate-pore gradient electrophoresis: comparison of tris-glycine and tris-borate-EDTA buffer systems. Anal Biochem. 1981 Jun;114(1):131–139. doi: 10.1016/0003-2697(81)90463-2. [DOI] [PubMed] [Google Scholar]
  23. Provost T. T., Arnett F. C., Reichlin M. Homozygous C2 deficiency, lupus erythematosus, and anti-Ro (SSA) antibodies. Arthritis Rheum. 1983 Oct;26(10):1279–1282. doi: 10.1002/art.1780261017. [DOI] [PubMed] [Google Scholar]
  24. Reddy R., Tan E. M., Henning D., Nohga K., Busch H. Detection of a nucleolar 7-2 ribonucleoprotein and a cytoplasmic 8-2 ribonucleoprotein with autoantibodies from patients with scleroderma. J Biol Chem. 1983 Feb 10;258(3):1383–1386. [PubMed] [Google Scholar]
  25. Rijken D. C., Collen D. Purification and characterization of the plasminogen activator secreted by human melanoma cells in culture. J Biol Chem. 1981 Jul 10;256(13):7035–7041. [PubMed] [Google Scholar]
  26. Scopelitis E., Biundo J. J., Jr, Alspaugh M. A. Anti-SS-A antibody and other antinuclear antibodies in systemic lupus erythematosus. Arthritis Rheum. 1980 Mar;23(3):287–293. doi: 10.1002/art.1780230304. [DOI] [PubMed] [Google Scholar]
  27. Somerville L. L., Wang K. The ultrasensitive silver "protein" stain also detects nanograms of nucleic acids. Biochem Biophys Res Commun. 1981 Sep 16;102(1):53–58. doi: 10.1016/0006-291x(81)91487-x. [DOI] [PubMed] [Google Scholar]
  28. Sontheimer R. D., Maddison P. J., Reichlin M., Jordon R. E., Stastny P., Gilliam J. N. Serologic and HLA associations in subacute cutaneous lupus erythematosus, a clinical subset of lupus erythematosus. Ann Intern Med. 1982 Nov;97(5):664–671. doi: 10.7326/0003-4819-97-5-664. [DOI] [PubMed] [Google Scholar]
  29. Tan E. M., Cohen A. S., Fries J. F., Masi A. T., McShane D. J., Rothfield N. F., Schaller J. G., Talal N., Winchester R. J. The 1982 revised criteria for the classification of systemic lupus erythematosus. Arthritis Rheum. 1982 Nov;25(11):1271–1277. doi: 10.1002/art.1780251101. [DOI] [PubMed] [Google Scholar]
  30. Terhorst C., van Agthoven A., LeClair K., Snow P., Reinherz E., Schlossman S. Biochemical studies of the human thymocyte cell-surface antigens T6, T9 and T10. Cell. 1981 Mar;23(3):771–780. doi: 10.1016/0092-8674(81)90441-4. [DOI] [PubMed] [Google Scholar]
  31. 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]
  32. Venables P. J., Smith P. R., Maini R. N. Purification and characterization of the Sjögren's syndrome A and B antigens. Clin Exp Immunol. 1983 Dec;54(3):731–738. [PMC free article] [PubMed] [Google Scholar]
  33. Wolin S. L., Steitz J. A. Genes for two small cytoplasmic Ro RNAs are adjacent and appear to be single-copy in the human genome. Cell. 1983 Mar;32(3):735–744. doi: 10.1016/0092-8674(83)90059-4. [DOI] [PubMed] [Google Scholar]

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