Download PDF
Horm Metab Res 2001; 33(2): 115-120
DOI: 10.1055/s-2001-12401
Original Clinical

© Georg Thieme Verlag Stuttgart · New York

Sensitive Assay to Detect Thyroid Stimulating Antibody (TSAb) in the Presence of Thyroid Stimulation Blocking Antibody (TSBAb) in Serum

Y. Ochi 1 , K. Yamashiro 2 , N. Takasu 2 , Y. Kajita 1 , Y. Sato 3 , A. Nagata 3
  • 1 Research Institute for Production Development, Morimoto-cho, Shimogamo, Sakyo-ku, Kyoto, Japan
  • 2 Department of Internal Medicine, Univ. of Rhukyus, Nishihara, Okinawa, Japan
  • 3 Yamasa-Shoyu, Ltd., Choshi, Chiba, Japan
Further Information

Publication History

Publication Date:
31 December 2001 (online)

Also available at
    Permissions and Reprints

The detection of thyroid stimulating antibody (TSAb) activity in the presence of thyroid stimulation blocking antibody (TSBAb) in Graves’ serum is difficult because TSBAb blocks TSAb activity. We recently demonstrated that polyethylene glycol (PEG) augments TSAb activity in porcine thyroid cells (PTC) assay. This PEG-induced augmentation makes it possible to develop a sensitive assay to detect TSAb in the presence of TSBAb. We studied the effects of PEG on TSAb- and TSBAb-activities in PTC using 4 different preparations of the samples; (1) crude IgG using PEG 22.5 % precipitated fraction (PF) from Graves’ serum (0.2 ml), (2) crude IgG using PEG 12.5 % PF, (3) serum (50 µl), and (4) serum (50 µl) in the presence of 5 % PEG (final). When the effects of PEG on TSAb activity using crude IgG were examined, PEG 22.5 % PF showed significantly higher TSAb activity than PEG 12.5 % PF as reported previously. The augmentative effect of PEG on TSAb activity was also observed by the addition of 5 % PEG to serum. We also demonstrated that PEG augmented TSAb-activities even in TSBAb-positive serum by two methods (crude IgG using PEG 22.5 % PF and the addition of 5 % PEG to serum). TSBAb activities were expressed by two calculation methods (A = [1 - (a - b)/(c - d) × 100] and B = [1 - (a - d)/(c - d) × 100], where a is cAMP produced in the presence of bTSH and patient’s IgG, b is cAMP produced in the presence of patient’s IgG, c is cAMP produced in the presence of bTSH and normal IgG, and d is cAMP produced in the presence of normal IgG). In the presence of TSAb, the values of A method were always higher than those of B method, since TSAb stimulated cAMP synthesis. We have developed two sensitive methods to detect TSAb even in the presence of TSBAb in serum using PEG; 1) incubation of crude IgG using PEG 22.5 % PF from serum (0.2 ml), and 2) co-incubation of 5 % PEG with test serum (50 µl).

Key words:

Thyroid Stimulating Antibody (TSAb) - Thyroid Stimulation Blocking Antibody (TSBAb) - cAMP - Graves’ Disease - Polyethylene Glycol (PEG)

References

  • 1 Zakarija M, McKenzie M. Immunoglobulin G inhibitor of thyroid-stimulating antibody is a cause of delay in the onset of neonatal Graves’ disease.  J Clin Invest. 1983;  72 1352-1356
    Search in Google Scholar
  • 2 Macchia E, Concetti R, Carone G, Borgoni F, Fenzi G F, Pinchera A. Demonstration of blocking immunoglobulin G, having a heterogenous behaviour, in sera of patients with Graves’ disease: Possible coexistence of different autoantibodies directed to the TSH Receptor.  Clin Endocrinol. 1988;  28 147-156
    PubMedSearch in Google Scholar
  • 3 Miyauchi A, Amino N, Tamaki H, Kuma K. Coexistence of thyroid stimulating and thyroid blocking antibodies in a patient with Graves’ disease who had transient hypothyroidism.  Amer J Med. 1988;  85 418-420
    PubMedSearch in Google Scholar
  • 4 Tamai H, Kasagi K, Takaichi Y, Takamatsu J, Komaki G, Matsubayashi S, Konishi J, Kuma K, Kumagai L, Nagataki S. Development of spontaneous hypothyroidism in patients with Graves’ disease treated with antithyroidal drugs: Clinical immunological and histological findings in 26 patients.  J Clin Endocrinol Metab. 1989;  69 49-53
    PubMedSearch in Google Scholar
  • 5 Morita T, Tamai H, Oshima A, Mukuta T, Fukata S, Kuma K, Kumagai L F, Nagataki S. The occurrence of thyrotropin binding inhibitory immunoglobulin and thyroid stimulating antibodies in patients with silent thyroiditis.  J Clin Endocrinol Metab. 1990;  71 1051-1055
    PubMedSearch in Google Scholar
  • 6 Takasu N, Yamada T, Sato A, Nakagawa M, Komiya I, Nakagawa Y, Asawa T. Graves’ disease following hypothyroidism due to Hashimoto’s disease: Studies of eight cases.  Clin Endocrinol. 1990;  33 687-698
    Search in Google Scholar
  • 7 Kasagi K, Hidaka A, Endo K, Miyamoto S, Takeuchi R, Misaki T, Sakahara H, Konishi J. Fluctuating thyroid function depending on the balance between stimulating and blocking types of TSH receptor antibodies: a case report.  Thyroid. 1993;  3 315-318
    PubMedSearch in Google Scholar
  • 8 Michelangeli Y P, Poon C, Topliss D J, Colman P G. Specific effect of radioiodine treatment on TSAb and TBAb levels in patients with Graves’ disease.  Thyroid. 1995;  5 171-176
    PubMedSearch in Google Scholar
  • 9 Inui T, Kouki T, Yamashiro K, Hachiya T, Ochi Y, Kajita Y, Sato Y, Nagata A. Increase of thyroid stimulating activity in Graves’ IgG by high polyethylene glycol concentrations using porcine thyroid cell assay.  Thyroid. 1998;  8 319-325
    PubMedSearch in Google Scholar
  • 10 Yamashiro K, Takasu N, Kouki T, Inui T, Ochi Y, Kajita Y, Sato Y, Nagata A. Augmentation of thyroid stimulating antibody-stimulated cyclic adenosine monophosphate response by polyethylene glycol, polyvinyl alcohol and dextran; high sensitive porcine thyroid cell assay thyroid-stimulating antibody assay.  Thyroid. 1998;  9 263-271
    PubMedSearch in Google Scholar
  • 11 Yamashiro K, Kouki T, Takasu N, Inui Y, Ochi Y, Kajita Y, Sato Y, Nagata A. Mechanism of the augmentative effect of high polyethylene glycol (PEG) concentrations on the tyroid stimulating activity in TSAb-IgG using porcine thyroid cell (PTC) assay.  Endocr Res. 1999;  25 67-76
    PubMedSearch in Google Scholar
  • 12 Ochi Y, Inui T, Kouki T, Yamashiro K, Takasu Y, Kajita Y, Sato Y, Nagata A. Clinical usefulness of TSAb assay with high polyethylene glycol concentrations.  Horm Res. 1999;  51 142-149
    CrossrefPubMedSearch in Google Scholar
  • 13 Kamijo K, Nagata A, Sato Y. Clinical significance of a sensitive assay for thyroid-stimulating antibodies in Graves’ disease using polyethylene glycol at high concentrations and porcine thyroid cells.  Endocr J. 1999;  46 397-403
    PubMedSearch in Google Scholar
  • 14 Kouki T, Inui T, Yamashiro K, Hachiya T, Ochi Y, Kajita Y, Takasu N, Sato A, Nagata A. Demonstration of fragments with thyroid stimulating activity from thyroid stimulation blocking antibodies-IgG molecules by papain digestion.  Clin Endocrinol. 1997;  47 693-698
    CrossrefPubMedSearch in Google Scholar
  • 15 Takeda K, Takamatsu J, Kasagi K, Sakane S, Ikegami Y, Isotani H, Majima T, Majima M, Kitaoka H, Iida Y, Ikekubo K, Konishi J, Mozai T. Development of hyperthyroidism following primary hypothyroidism: A case report with changes in thyroid-related antibodies.  Clin Endocrinol. 1988;  28 341-344
    PubMedSearch in Google Scholar
  • 16 Chiovato L, Vitti P, Bendinelli G, Santani F, Fiore E, Capaccioli A, Tonacchera M, Mammoli C, Ludgate M, Pinchera A. Detection of antibodies blocking thyreotropin effect using Chinese hamster ovary cells transfected with the cloned human TSH receptor.  J Endocrinol Invest. 1994;  17 809-816
    PubMedSearch in Google Scholar
  • 17 Kasagi K, Nagata A, Yoshimura H, Misaki T, Sato Y, Konishi J. Development and evaluation of a thyroid stimulation blocking antibody (TSBAb) assay using dispersed porcine thyroid cells. The proceedings of 71st Annual Meeting of the American Thyroid Association (abstract), 1998: 92
    Search in Google Scholar
  • 18 Konishi J, Iida Y, Endo K, Misaki T, Nohara Y, Matuura N, Mori T, Torizuka K. Inhibition of thyrotropin-induced adenine 3’5’-monophosphate increase by immunoglobulin from patients with primary myxedema.  J Clin Endocrinol Metab. 1983;  57 544-549
    PubMedSearch in Google Scholar
  • 19 Inomata H, Sasaki N, Tamaru K, Ushiku H, Nimi H, Nakajima H. Relationship between potency of blocking type thyrotropin-binding inhibitor immunoglobulin in three women with primary myxedema and thyroid function of their neonates.  Endocr Jap. 1986;  33 353-359
    PubMedSearch in Google Scholar
  • 20 Yokoyama N, Izumi M, Katamine S, Nagataki S. Heterogeneity of Graves’ immunoglobulin G; comparison of thyrotropin receptor antibodies in serum and in culture supernatants of lymphocytes transformed by Epstein-Barr virus infection.  J Clin Endocrinol Metab. 1987;  64 215-218
    PubMedSearch in Google Scholar
  • 21 Matsui I, Sakata S, Ogawa T, Takuno H, Sarui H, Komaki T, Manshouri T, Atassi Z. Biological activities of rat antisera raised against synthetic peptide of human thyrotropin receptor.  Endocr J. 1993;  40 607-612
    PubMedSearch in Google Scholar
  • 22 Hidaka Y, Guimaraes V, Soliman M, Yanagawa T, Okamoto Y, Quintans J, DeGroot L J. Production of thyroid-stimulating antibodies in mice by immunization with T-cell epitopes of human thyrotropin receptor.  Endocrinology. 1995;  136 1642-1647
    CrossrefPubMedSearch in Google Scholar
  • 23 Zhu Y, Portmann L, Denereaz L, Lemarchand-Beraud T. Simultaneous assay for three types of thyrotropin receptor antibody activities using FRTL-5 cells in patients with autoimmune thyroid disease.  Eur J Endocrinol. 1994;  131 359-368
    PubMedSearch in Google Scholar
  • 24 Ochi Y, Yoshimura M, Hachiya T, Miyazaki T. Immunological studies on LATS-immunoglobulin by the reaction with Staphylococcal Protein A.  Endocrinol J. 1976;  23 183-186
    PubMedSearch in Google Scholar
  • 25 Morgenthaler N G. New assay systems for thyrotropin receptor antibodies.  Curr Opin Endocrinol Diabetes. 1999;  6 251-260
    CrossrefPubMedSearch in Google Scholar

Y. Ochi,M.D. 

Research Institute for Production Development

15 Morimoto-cho, Shimogamo
Sakyo-ku, Kyoto 6060805
Japan


Phone: Phone:+ 81 (075) 781-1107

Fax: Fax:+ 81 (075) 791-7659