Clinical Chemistry Link to Randox Laboratories Web Site
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Clinical Chemistry 43: 1392-1396, 1997;
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Submit an electronic Letter to
the Editor about this paper
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via ISI Web of Science (5)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Kaczur, V.
Right arrow Articles by Balázs, Cs.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Kaczur, V.
Right arrow Articles by Balázs, Cs.
Related Collections
Right arrow Clinical Immunology
Right arrow Endocrinology and Metabolism
(Clinical Chemistry. 1997;43:1392-1396.)
© 1997 American Association for Clinical Chemistry, Inc.

Articles

Effect of anti-thyroid peroxidase (TPO) antibodies on TPO activity measured by chemiluminescence assay

V. Kaczur1,a, Gy. Vereb2, I. Molnár1, G. Krajczár1, E. Kiss1, N. R. Farid3 and Cs. Balázs1

1 Department of Pathology, III Department of Medicine, Kenézy Teaching Hospital, Bartók B. 2–26, H-4043 Debrecen, Hungary. Fax 36 (52) 418 880; e-mail stefan{at}tigris.klte.hu

2 Department of Medical Chemistry, University Medical School of Debrecen, Hungary. Fax 36 (52) 412 566; e-mail vereb{at}dragon.klte.hu

3 Internal Medicine-Endocrinology and Metabolism, Chatham, Ontario, Canada. Fax (519) 351 8680; e-mail nfarid{at}wwdc.com
a Author for correspondence.


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
A chemiluminescence method was developed to measure thyroid peroxidase (TPO) activity and the inhibitory effect of anti-TPO antibodies in purified porcine TPO. The TPO preparation was characterized kinetically and controlled by Western-blotting technique. The chemiluminescence method proved to be reproducible and much more sensitive than the widely used guaiacol method, being able to detect TPO concentrations of 2.21 x 10-5 g/L vs 6.63 x 10-2g/L with the latter. Otherwise, the determinations with the two methods correlated well (r = 0.76). Investigating the effect of IgGs from 23 hypothyroid patients on measured TPO activity, we detected inhibition in 19 cases with the chemiluminescence technique (15 with the guaiacol method). Anti-TPO antibodies showed competitive inhibition of TPO activity with respect to the substrate guaiacol. In both systems, the inhibition is present in the IgG F(ab')2 fragment. We conclude that the high sensitivity of chemiluminescence detection allows routine determination of the inhibition of TPO activity by anti-TPO antibodies.


Key Words: indexing terms: kinetic characterization • immunoinhibition • autoimmune disease • thyroiditis


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
Thyroid peroxidase (TPO) is a major thyroid microsomal antigen (M/TPO) (1)(2) in autoimmune thyroiditis (3)(4). A glycoprotein (5)(6)(7) sensitive to reductive agents (8), TPO is an integral part of thyroid membrane and plays a key role in the iodination of tyrosyl residues in thyroglobulin and other proteins (9)(10)(11)(12). Evaluation of the diagnostic value of an anti-TPO assay in patients with different types of thyroid disease and in controls showed increased anti-TPO antibodies in >90% patients with active autoimmune thyroiditis (13)(14). The antigenic determinants of TPO-binding anti-TPO antibodies are heterogeneous (largely of IgG1 and IgG4 subclasses) (15). The antibodies are produced in both thyroidal and extrathyroidal compartments (15).

The autoimmune response to TPO is known to be associated with a large number of different epitopes on TPO, including the enzymatic site (11)(16), and two methods accepted for determination of TPO are based on oxidation of iodide (17) or guaiacol [9]. However, these methods require relatively high amounts of purified TPO. The inhibition of TPO by anti-TPO antibodies varies with the method used (iodide or guaiacol), because the inhibition of TPO activities measured by the assays can be caused by autoantibodies that recognize either one of the enzyme's binding sites (18). Other polyclonal anti-TPO antibodies are assumed not to bind to the catalytic site of molecule because they do not modify the activity of enzyme (19). The aims of the present study were: (a) to develop a new chemiluminescence technique for determination of TPO activity, (b) to obtain purified TPO antigen suitable for testing the effect of anti-TPO IgGs on enzyme activity, (c) to test the possible inhibitory activity of anti-TPO antibodies from patients with Hashimoto thyroiditis, and (d) to study the kinetics of the inhibition.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
preparation of porcine tpo
This was performed essentially according to Neary et al. (9). Porcine thyroid glands were ground and washed four times by suspending in 1.15 mol/L KCl plus 0.1 mmol/L KI and centrifuging (5000g, 40 min). The tissue was then homogenized with 0.25 mol/L sucrose plus 0.1 mmol/L KI and filtered through cheesecloth; the filtrate was centrifuged (5000g, 20 min). From the supernatant the microsome fraction was pelleted by ultracentrifugation at 50 000g for 180 min and washed successively with (a) 1 mol/L NaCl, 10 mmol/L Tris-HCl, and 0.1 mmol/L KI, pH 7.4, and (b) 0.1 mmol/L KI. The washed microsome fraction was treated with chymotrypsin-free, diphenylcarbamyl chloride-treated trypsin (35 kU/g protein; Merck, Darmstadt, Germany) at 37 °C for 1 h and the reaction was terminated with soybean trypsin inhibitor (2 mg/mg trypsin). The supernatant obtained after 105 000g for 1 h was applied to an 1.6 x 22 cm anion-exchange column (DE-52; Whatman, Maidstone, Kent, UK) equilibrated with a solution of 25 mmol/L KCl, 0.1 mmol/L KI, and 15 mmol/L Tris-HCl, pH 7.4. The TPO activity was eluted by applying a linear KCl gradient (from 25 to 225 mmol/L KCl) in 0.1 mmol/L KI plus 15 mmol/L Tris-HCl, pH 7.4. The pooled fractions were concentrated with a PM-30 ultrafiltration membrane (Amicon, Oosterhout, The Netherlands), applied to a 1.6 x 85 cm Sephacryl S-200 (Pharmacia, Uppsala, Sweden) column equilibrated with 50 mmol/L potassium phosphate plus 0.1 mmol/L KI, pH 7.2, and eluted with the same buffer. The active fractions were concentrated as before and stored frozen at -70 °C.

identification of purified tpo fragments
The activity of purified porcine TPO was measured after all steps (Table 1 ), and the material was subjected to sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) and monitored by Western blotting. SDS-PAGE was carried out by the method of Laemmli (20), with use of a 80 g/L separating gel in conjunction with a 100 g/L stacking gel. The prepared TPO fractions and prestained molecular mass markers (MW SDS 7B; Sigma, St. Louis, MO), 0.15 mL each, were boiled for 5 min in 0.3 mL of sample-reducing buffer (Reanal, Budapest, Hungary) consisting of 20 g/L SDS, 50 mL/L 2-mercaptoethanol, 100 mL/L glycerol, and 65 mmol/L Tris and were electrophoresed at 30–40 mA for 10 h. Separated gel proteins were stained with Amidoblack 10B (Sigma).


View this table:
[in this window]
[in a new window]
  		Table 1. Purification of TPO from frozen porcine thyroid tissue.

Western blotting was performed as described by Towbin et al. (21). The purified TPO proteins were electrotransferred onto nitrocellulose filters (0.45 nm pore size; Paraplan, Budapest, Hungary) at 180 mA for 24 h at 4 °C. Filters were incubated in Cohne-buffer (Reanal) consisting of 1 g/L Ficoll, 1 mL/L Tween 20, 15 g/L bovine serum albumin, 0.5 g/L gelatin, 0.02 mol/L Tris, and 100 mL/L glycerol and were cut into 3-mm-wide strips. The strips, in groups of three, were incubated overnight with 3 mL of 40-fold-diluted serum samples at room temperature. After washing, the strips were incubated at room temperature for 1 h with 1000-fold-diluted goat anti-human IgG and IgA antibodies conjugated with horseradish peroxidase (Sigma). The strips were stained at room temperature with 0.2 g/L diaminobenzidine (Serva, Heidelberg, Germany) in 1 mol/L Tris-buffered saline (pH 7.4) containing 20 µL of 300 mL/L hydrogen peroxide solution.

measurement of tpo activity
Guaiacol method.
TPO activity was routinely determined in a 1.0-cm-lightpath cuvette containing 33 mmol/L guaiacol (Fluka, Buchs, Germany) in a 1-mL solution of 0.1 mol/L phosphate buffer, pH 7.4, and 0.3 mmol/L hydrogenperoxide (Merck). The reaction was initiated by adding H2O2. The rate of reaction was calculated with an absorptivity value of 5.75 L cm-1 mmol-1 for oxidized guaiacol (9); 1 G.U. was defined as the amount of enzyme that transformed 1 µmol of guaiacol per minute. For kinetic measurements the guaiacol concentrations were varied between 0.5 and 48 mmol/L and the H2O2 concentrations between 0.3 and 0.85 mmol/L.

Peroxidase activity was assayed only by the guaiacol method, because Hosoya and Morrison had found in 1967 that the purified porcine TPO could oxidize 48.9 µmol of guaiacol per min/mg protein, but only 7.35 µmol of iodide ion per min/mg protein (1).

Chemiluminescence method
. This method is based on the oxidation of luminol by H2O2 as catalyzed by the peroxidase (22). Chemiluminescence was measured with a Luminometer 1250 (LKB Wallac, Uppsala, Sweden). The 1-mL reaction mixture contained 1 mol/L glycine–NaOH buffer, pH 9, 1 mmol/L EDTA, 10 µmol/L luminol, and aliquots of TPO. The reaction was initiated by injection of 0.1 mmol/L H2O2; the emitted signals were measured at 37 °C. The resulting light output was determined in mV with a chart recorder (LKB 2210). The luminol–peroxidase blank was simultaneously determined in every measurement.

igg isotype of anti-tpo antibodies
IgG was obtained from sera of patients or healthy controls by precipitation with polyethylene-glycol Mr 6000 (PEG 6000; Fluka), as reported by Tamai et al. (23). The IgG isotype anti-TPO antibodies were separated by affinity chromatography on a Protein G–Sepharose (Pierce, Rockford, IL) column. The eluted antibodies were desalted by gel-filtration. IgG fragments were obtained by treating the IgG solution with pepsin (Pierce) and the resulting IgG Fc and IgG F(ab')2 fragments were separated on a Protein A–Sepharose column (also from Pierce) (24)(25). Elution of the IgG was monitored at 280 nm.

inhibition of tpo activity by antibodies
In the guaiacol system (kinetic method), we in general used 3.4 µmol/L IgG (in the kinetic measurements, 1.8–105 µmol/L), Mr 156 000, after separation by PEG precipitation, and 3.7 g/L IgG F(ab')2. The solution containing anti-TPO antibodies was incubated with TPO with shaking at 23 or 37 °C. Control IgGs (from healthy subjects) were also incubated the same way. The results were corrected for the absorbance of IgG alone in every measurement.

In the luminol system (nonkinetic method), we routinely preincubated TPO with 1.78 pmol/L PEG-precipitated IgG (in studies, 0.178–89.1 pmol/L IgG) and 5.4 mg/L–1.54 g/L IgG F(ab')2 used as above.

In routine measurements, the degree of inhibition is expressed as "inhibition percent" in both systems.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Purified TPO showed immunoreactivity at 105 and 110 kDa under nonreducing conditions (data not shown). The specific activity of the final porcine TPO preparation determined with the guaiacol method was 14.18 kG.U./g protein, similar to the value reported Neary et al. (9). The Km for guaiacol determined according to Lineweaver–Burk was 5.6 x 10-4 mol/L (26); Km values of 10-4 mol/L, 3 x 10-6 mol/L, and 6 x 10-3 mol/L were found for protein iodination (5), Glu-Tyr-Glu iodination (6), and iodide-oxidation (7), respectively.

In a previous luminescent horseradish peroxidase assay, a relationship was demonstrated between the concentrations of the peroxidase and the intensity of light emitted (27). Our assay, based on luminol-dependent chemiluminescence, detected TPO concentrations between 22.1 µg/L and 66.3 mg/L (the mean + 3SD signal of zero calibrator is <=22.1 µg/L) (Fig. 1 ).



View larger version (14K):
[in this window]
[in a new window]
  		Figure 1. Chemiluminescence assay of TPO activity: TPO dependency of chemiluminescence.

TPO concentration ranged from 66 mg/L to 22 µg/L; initial luminol concentration was kept at 10 µmol/L. Assay performed as described in Materials and Methods.

Inhibition of TPO activity by anti-TPO antibodies was proportional to antibody concentrations in both systems. In the chemiluminescence method this inhibition was detected in the range 0.178–89.1 pmol/L for IgG (Fig. 2 ) and 27.7 mg/L to 1.54 g/L for IgG F(ab')2 (Fig. 3 ). Interassay variation (CV) was 11.3%. To characterize the type of inhibition involved, we examined various substrate and inhibitor concentrations and found a competitive inhibition of IgG isotype anti-TPO antibodies with guaiacol substrate (26). The inhibition coefficient was determined by Dixon's diagram as 20.1 µmol/L for IgG (Fig. 4 ; see also our previous study (26)). The inhibition can be attributed to F(ab')2 fragment (in both systems) (26). A few workers reported an effect of anti-TPO antibodies binding on enzyme activities, but only one part of anti-TPO antibodies was able to inhibit the TPO activity (18)(28). In our experiment, IgGs from 15 of the 23 patients with Hashimoto disease inhibited the TPO activity in the guaiacol method; in contrast, 19 showed TPO inhibition in the chemiluminescence technique. Agreement between the determinations performed by the chemiluminescence and guaiacol methods was modest (Fig. 5 ).



View larger version (13K):
[in this window]
[in a new window]
  		Figure 2. Effect of anti-TPO antibodies on porcine TPO activity determined by chemiluminescence method.

IgG isotype anti-TPO antibodies were obtained by PEG precipitation; inhibition as described in text.



View larger version (13K):
[in this window]
[in a new window]
  		Figure 3. Inhibition of porcine TPO by IgG F(ab')2 fragments of anti-TPO antibodies measured by chemiluminescence method.

Preparation of IgG F(ab')2 fragments of antibodies and the range of inhibition were as described in the text.



View larger version (14K):
[in this window]
[in a new window]
  		Figure 4. Inhibition of porcine TPO activity by IgG isotype of anti-TPO antibodies, according to Dixon.

The inhibitor coefficient was determined as 2.3 x 10-5 mol/L by Dixon's diagram, representing 1/v as a function of [I] for different [S]: *, 12 mmol/L; •, 20 mmol/L; +, 33 mmol/L. Composition of the reaction mixture was as given in the text (v was calculated as nanomoles of guaiacol oxidized per minute per milligram of protein).



View larger version (14K):
[in this window]
[in a new window]
  		Figure 5. Comparison of guaiacol and chemiluminescence methods for inhibition of TPO.

The degree of inhibition was calculated as "inhibition percent" in both systems: guaiacol % (y) and luminol % (x); y = 0.75x + 15.8 (r = 0.76).


   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
Evidence has accumulated in the last few years that anti-TPO antibodies are closely associated with the activity of autoimmune thyroiditis (29). These antibodies are believed to be of greater pathogenetic significance than anti-thyroglobulin antibodies in thyroid cellular dysfunction and in the pathogenesis of hypothyroidism (30). Anti-TPO antibodies fix complement, resulting in destruction of thyroid gland (31). Measurements of anti-TPO antibodies are used for diagnosis and determination of disease activity of thyroiditis (3). The observation concerning the capability of one type of anti-TPO antibodies to inhibit the enzyme activity has suggested a new mechanism that might be involved in thyroiditis. The functional determination of anti-thyroid antibodies, including anti-TPO, is thus potentially important in autoimmune thyroid patients. The conventional guaiacol and iodide techniques require relative large amounts of TPO and are not used routinely. The chemiluminescence method developed for measuring TPO activity has several advantages over other methods. It is more sensitive than the usual guaiacol method (detecting as little TPO as 22.1 µg/L vs only 66.3 mg/L in the guaiacol method) and requires much less TPO preparation for the detection of IgG inhibition. Also, its specificity and reproducibility make it suitable for testing a relatively large number of samples, thus making the method potentially more economical.

Although in vitro a substantial proportion of anti-TPO antibodies inhibit the activity of this enzyme, it is not clear whether in vivo these antibodies mediate thyroiditis by regulating or modifying the TPO activity. Our observations support the concept of an inhibitory effect of anti-TPO on thyroid hormone production, in that 15 or 19 IgGs from 23 patients with hypothyroidism inhibited TPO activity. A direct effect on the enzyme activity is supported by the finding that anti-TPO antibodies exert competitive inhibition with the substrate guaiacol (26). Our prospective study using the chemiluminescence method should provide further evidence on the possible pathogenetic role of inhibitory-type anti-TPO antibodies in autoimmune thyroid disease.


   Acknowledgments
 
This work was supported by grants from the Academy of Hungarian Sciences (OTKA T 020913 and ETT 023/93).


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Czarnocka B, Ruf J, Ferrand M, Carayon P, Lissitzky S. Purification of the human thyroid peroxidase and its identification as the microsomal antigen involved in autoimmune thyroid disease. FEBS Lett 1985;190:147-152. [ISI][Medline] [Order article via Infotrieve]
  2. Portmann L, Hamada N, Heinrich G, Degroot LJ. Anti-thyroid peroxidase antibody in patients with autoimmune thyroid disease: possible identity with anti-microsomal antibody. J Clin Endocrinol Metab 1985;61:1001-1003. [Abstract]
  3. Chiovato L, Pinchera A. The microsomal-peroxidase antigen: modulation of its expression in thyroid cells. Autoimmunity 1991;10:319-331. [ISI][Medline] [Order article via Infotrieve]
  4. Czarnocka B, Ruf J, Ferrand M, Carayon P. The immunochemical properties of human thyroid peroxidase. Thyroperoxidase Thyr Autoimm 1990;201:59-67.
  5. Rawitch AB, Taurog A, Chernoff SB, Dorris ML. Hog thyroid peroxidase: physical, chemical, and catalytic properties of the highly purified enzyme. Arch Biochem Biophys 1979;194:244-257. [ISI][Medline] [Order article via Infotrieve]
  6. Neary JT, Koespell D, Davidson B, Armstrong A, Strout HV, Soodak M, Maloof F. Interaction of thyroid peroxidase with concanavalin A covalently coupled to agarose. J Biol Chem 1977;252:1264-1271. [Abstract/Free Full Text]
  7. Davidson B, Neary JT, Strout HV, Maloof F, Soodak M. Evidence for a thyroid peroxidase associated "active iodine" species. Biochim Biophys Acta 1978;522:318-326. [Medline] [Order article via Infotrieve]
  8. Gardas A, Domek H. The effect of sulphydryl reagents on the human thyroid microsomal antigen. J Endocrinol Invest 1988;11:385-388. [ISI][Medline] [Order article via Infotrieve]
  9. Neary JT, Soodak M, Maloof F. Iodination by thyroid peroxidase [Review]. Methods Enzymol 1984;107:445-475. [ISI][Medline] [Order article via Infotrieve]
  10. Nunez J, Pommier J. Formation of thyroid hormones. Vitam Horm 1982;39:175-229. [ISI][Medline] [Order article via Infotrieve]
  11. Banga JP, McGregor AM. Enzymes as targets for autoantibodies in human autoimmune disease: relevance to pathogenesis. Autoimmunity 1991;9:177-182. [ISI][Medline] [Order article via Infotrieve]
  12. Ekholm R. Biosynthesis of thyroid hormones. Int Rev Cytol 1990;120:243-288. [ISI][Medline] [Order article via Infotrieve]
  13. Engler H, Riesen WF, Keller B. Diagnostic value of autoantibodies against microsomal thyroid peroxidase (anti-TPO). Schweiz Med Wochenschr 1992;122:1976-1980. [ISI][Medline] [Order article via Infotrieve]
  14. Laurberg P, Pedersen KM, Vittinghus E, Ekelund S. Sensitive enzyme-linked immunosorbent assay for measurement of autoantibodies to human thyroid peroxidase. Scand J Clin Lab Invest 1992;52:663-669. [ISI][Medline] [Order article via Infotrieve]
  15. DeGroot LJ. Heterogeneity of human autoantibodies to TPO. Thyroperoxidase Thyr Autoimm 1990;207:177-182.
  16. Ruf J, Toubert ME, Czarnocka B, Durande-Gorde JM, Ferrand M, Carayon P. Relationship between immunological structure and biochemical properties of human thyroid peroxidase. Endocrinology 1989;125:1211-1218. [Abstract]
  17. Hosoya T, Morrison M. The isolation and purification of thyroid peroxidase. J Biol Chem 1967;242:2828-2833. [Abstract/Free Full Text]
  18. Kohno Y, Hiyama Y, Shimojo N, Niimi H, Nakajima H, Hosoya T. Autoantibodies to thyroid peroxidase in patients with chronic thyroiditis effect of antibody binding on enzyme activities. Clin Exp Immunol 1986;65:534-541. [ISI][Medline] [Order article via Infotrieve]
  19. Venot N, Vinet C, Carayon P. Autoantibodies to TG and/or TPO do not inhibit thyroid hormone genesis. J Endocrinol Invest 1994;17:26.
  20. Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 1970;227:680-685. [Medline] [Order article via Infotrieve]
  21. 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;76:4350-4354. [Abstract/Free Full Text]
  22. Whitehead TP, Kricka LJ, Carter TJN, Thorpe GHG. Analytical luminescence: its potential in the clinical laboratory [Review]. Clin Chem 1979;25:1531-1546. [Abstract/Free Full Text]
  23. Tamai H, Amino N, Iwatani Y, Aozasa M, Kanay S, Nose S, et al. Highly sensitive bioassay for TSH receptor antibodies using FRTL-5 cells. Clin Endocrinol 1986;9:814.
  24. Arkerstorm B, Bjorck L. A physicochemical study of Protein G, a molecule with unique immunoglobulin G-binding properties. J Biol Chem 1986;261:10240-10247. [Abstract/Free Full Text]
  25. Arkerstorm B. Protein G. A powerful tool for binding and detection of monoclonal and polyclonal antibodies. J Immunol 1985;135:2589-2592. [Abstract]
  26. Kaczur V, Vereb Gy, Molnár I, Krajczár G, Balázs Cs. Kinetic characterization of thyroid peroxidase enzyme and its inhibition by autoantibodies. Acta Microbiol Immunol Hung 1995;42:345-350. [Medline] [Order article via Infotrieve]
  27. Puget K, Michelson AM, Avrameas S. Light emission techniques for the microestimation of femtogram levels of peroxidase: application to peroxidase (and other enzymes)-coupled antibody–cell antigen interactions. Anal Biochem 1977;79:447-456. [ISI][Medline] [Order article via Infotrieve]
  28. Okamoto Y, Hamada N, Saito H, Ohno M, Noh J, Ito K, Morii H. Thyroid peroxidase activity-inhibiting immunoglobulins in patients with autoimmune thyroid disease. J Clin Endocrinol Metab 1989;68:730-734. [Abstract]
  29. Rapoport B. Pathophysiology of Hashimoto-thyroiditis and hypothyrodism. Annu Rev Med 1991;42:91-96. [ISI][Medline] [Order article via Infotrieve]
  30. Weetman AP, McGregor AM. Autoimmmune thyroid disease: further developments in our understanding. Endocr Rev 1994;15:788-814. [Abstract]
  31. Chivato L, Bassi P, Santini F, Mammoli C, Lapi P, Carayon P, Pinchera A. Antibodies producing complement-mediated thyroid cytotoxicity in patients with atrophic or goitrous autoimmune thyroiditis. J Clin Endocrinol Metab 1993;77:1700-1705. [Abstract]




This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Submit an electronic Letter to
the Editor about this paper
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via ISI Web of Science (5)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Kaczur, V.
Right arrow Articles by Balázs, Cs.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Kaczur, V.
Right arrow Articles by Balázs, Cs.
Related Collections
Right arrow Clinical Immunology
Right arrow Endocrinology and Metabolism

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS