A new assay for thyroglobulin concentration in serum using monoclonal antibodies against synthetic peptides

The concentration of thyroglobulin (Tg) measured by radioimmunoassay (RIA) or enzyme-linked immunosorbent assay (ELISA) is greatly affected by the presence of anti-Tg autoantibodies in sera. We developed a new assay for detecting Tg in the presence of high concentrations of anti-Tg autoantibodies. A 48-kDa fragment was purified from Tg after treatment with V8 protease. This fragment did not appear to bind to two types of monoclonal antibodies (57Ab and 28D3) against a peptide in the C-terminus (amino acids 2735-2748) of Tg and intact Tg, respectively, by ELISA and Western blot analysis. In contrast, anti-Tg autoantibody or anti-Tg polyclonal antibody reacted well with this fragment. Our new ELISA used 57Ab as a solid phase antibody and 28D3 as a antibody conjugated to horseradish peroxidase. Buffer containing purified 48-kDa fragment was used to neutralize autoantibodies against Tg. With this assay, the recovery of Tg was 84.0-89.6% in normal healthy donors (n=5) in the presence of immunoglobulin G (IgG) purified from sera positive for anti-Tg autoantibody, and 76.2-104.4% in patient sera Grave's disease (n=15). Furthermore, the Tg concentrations in sera from patients with Grave's disease (n=20) ranged from 25 to 526 ng/ml, even though the Tg concentration, as measured by a commercial RIA did not exceed 55 ng/ml. There was good agreement between Tg concentrations measured by new Tg-ELISA and commercial Tg-RIA in sera that were negative for anti-Tg autoantibody. Overall, our new ELISA containing a Tg fragment to neutralize the presence of autoantibodies, showed good sensitivity and precision, and may be useful for routine use. Further investigations with the new assay should allow wider assessment of the prevalence and pattern of thyroid autoimmunity or thyroid neoplasms.     

Demonstration of circulating autoantibodies against the plasma membrane of isolated thyroid cells in Graves' disease

Antibodies against the plasma membrane of isolated human and porcine thyroid cells were demonstrable by a linear fluorescence pattern in sera from 29 out of 37 patients with Graves' disease. In 3 cases the antibodies reacted only with human thyroid cells. In a group of patients with Hashimoto's thyroiditis, 6 of 8 sera reacted with human thyroid cells in a granular fluorescence pattern. With one exception, sera from patients with non-toxic goiter showed no binding of IgG to thyroid cells. Sera from healthy controls were negative in this test system. There was no correlation between the presence of antibodies against thyroid cells and antibodies against thyroglobulin. Antibodies against thyroid microsomal antigens were found in 80% of patients with circulating thyroid cell membrane antibodies; 5 sera with membrane antibodies were negative for microsomal antibodies. Twenty sera of patients with Graves' disease were tested by passive haemagglutination test against peak I and peak II of the human thyroid fractionation with Sephadex G100. Antibodies against peak I were found in 14 sera and against peak II in all 20 sera. The data indicate that the antibodies in sera from patients with Graves' disease are directed mainly against a species-unspecific antigen of the plasma membrane from thyroid cells with a probable molecular weight between 4-7 S and are unrelated to thyroglobulin.     

The role of self-antigen in the development of autoimmunity in Obese strain chickens with spontaneous autoallergic thyroiditis

Neonatal thyroidectomy of Obese strain (OS) chickens showed that the spontaneous development of thyroid autoimmunity in these animals was fully dependent upon the presence of autoantigen, and could not be ascribed essentially to antigen-independent mechanisms such as polyclonal lymphocyte activation or innate distortions within the idiotype network. Similarly, removal of the gland in animals with established thyroiditis demonstrated the need for antigen to maintain the autoimmune response. Thyroglobulin from normal chickens induced autoantibodies in neonatally thyroidectomized OS birds, suggesting that an abnormality in the structure of this protein is not a prerequisite for the development of autoimmunity. This contention is supported by the finding that OS and normal thyroglobulin were immunochemically indistinguishable, whether compared using OS autoantibodies or rabbit anti-chicken thyroglobulin sera.     

On the presence of autoantibodies in the plasma of patients suffering from medullary carcinoma of the thyroid

Thyroiditis is associated with certain cases of medullary carcinoma of the thyroid (MCT). In order to determine if this was due to the presence of autoantibodies, we have investigated whether plasma from patients suffering from the disease contain autoantibodies. The specificity of the reaction was established using specific antibodies to antigens such as: calcitonin (CT), carcinoembryonic antigen (CEA) and thyroglobulin (Tg); no antibodies against CT, CEA or Tg were detected in the plasma from the patients investigated. The presence of autoantibodies to a constituent of the tumor was established in each case studied. The detection of autoantibodies to a "specific" constituent of MCT could be of potential interest in the early screening for the disease.     

Detection of antithyroglobulin autoantibodies with defined epitopic specificity by a microparticle-enhanced nephelometric immunoassay.

To hydrophilic, polyfunctional spherical microparticles of predetermined diameter, produced by copolymerization of acrylic monomers, we covalently bound human thyroglobulin. The thyroglobulin-microsphere conjugate was agglutinated, in the presence of antimouse immunoglobulins antiserum, by four monoclonal antibodies, each recognizing a different antigenic domain on the thyroglobulin molecule. These agglutinations were quantified by measuring with a specially designed nephelometer the light scattered by clusters of the conjugates. Agglutination with the monoclonal antibody recognizing antigenic domain II of the thyroglobulin molecule was specifically inhibited by some human sera that contained antithyroglobulin autoantibodies. This allowed us to develop a microparticle-enhanced nephelometric immunoassay for these autoantibodies with defined epitopic specificity. Using this assay, we detected and quantified antithyroglobulin autoantibodies in serum samples from all eight patients examined with Hashimoto disease and from most (75%) patients with untreated Graves disease.     

Radioimmunoassay for Measurement of Thyroglobulin in Human Serum

A specific and reproducible double antibody radioimmunoassay for the measurement of thyroglobulin (HTg) in human serum has been developed. Since antithyroglobulin autoantibodies combine with the [(131)I] HTg tracer, antibody-positive sera were rejected for measurement. Specificity is demonstrated in that thyroid analogous such as thyroxine (T(4)), triiodothyronine (T(2)) monoiodotyrosine (MIT) and diiodotyrosine (DIT) did not crossreact. Sera previously reacted with anti-HTg-Sepharose contained no immunoassayable HTg. Finally, sera obtained from patients after total thyroid ablation for thyroid carcinoma did not contain demonstrable HTg. The sensitivity of the assay is 1.6 ng/ml, and HTg was detectable in 74% of 95 normal subjects. The mean concentration was 5.1 ng/ml +/-0.49 SEM (range <1.6-20.7 ng/ml). Day to day variation in HTg levels is large in some euthyroid subjects and nearly absent in others. HTg was detectable in 90% of the sera obtained in 23 pregnant women at delivery in whom a mean concentration of 10.1 ng/ml +/-1.3 SEM was observed. The mean level for the corresponding newborn infants at birth was 29.3 ng/ml +/-4.7 SEM a value significantly higher than the mean maternal HTg concentration (P <0.01). A group of 17 thyrotoxic individuals all had elevated HTg levels; the mean for this group was 344.8 ng/ml +/-90.7 SEM. In the acute phase of subacute thyroiditis HTg was also elevated in all of 12 patients, and the mean for this group was 136.8 ng/ml +/-74.6 SEM.     

Antigen binding characteristics of thyroid microsomal autoantibodies studied with ELISA methods

A competitive fluorometric ELISA was developed which enabled pair-wise comparisons of the antigen-binding characteristics of thyroid microsomal autoantibodies (TMaab) and their antigen-binding fragments from different individuals. Antibodies rendered undetectable to the protein-A-enzyme conjugate by removal of their Fc-regions (F(ab')2's) were tested for their ability to inhibit the antigen-binding of intact thyroid microsomal autoantibodies (TMaab). Only F(ab')2's derived from TMaab-containing sera were capable of displacing TMaab, some being capable of complete displacement of allogeneic TMaab. The antigen-binding of Tgaab was unaffected, except by F(ab')2's derived from Tgaab-containing sera. The high degree of overlap observed in the specificity of TMaab suggests that different sera recognize the same epitopes. In this respect the autoimmune response to the microsomal antigen was shown to resemble that to thyroglobulin, which involves few epitopes. Cross-tolerance is postulated to be the mechanism underlying the tissue-specificity and epitope-restriction of thyroid autoantibodies.     

Elevated serum thyroglobulin. A marker of metastases in differentiated thyroid carcinomas.

The presence of human thyroglobulin (HTg) in serum of patients was identical by immunological criteria to the serum standard used in the radioimmunoassay. The serum thyroglobulin levels in untreated patients with differentiated thyroid carcinoma ranged from 22.0 to 445.0 ng/ml with a mean of 144.3 +/- 46.5 ng/ml (SEM) (n = 10). The mean serum thyroglobulin measured postoperatively in seven of these patients was 6.4 +/- 1.5 ng/ml, not statistacally different from the mean level of 5.1 +/- 0.49 ng/ml (range 0-20.7 ng/ml) observed in 71 out of 95 control subjects with detectable HTg levels. By contrast serum HTg levels were normal or undetectable in subjects with medullary carcinoma of the thyroid. HTg levels were within normal limits in sera of patients who had previously undergone successful therapy for a differentiated thyroid carcinoma and in whom no metastases could be documented. The mean level for this group was 4.9 +/- 0.51 ng/ml (n = 43). In contrast, patients with documented metastases had a mean serum thyroglobulin level of 464.9 +/- 155.6 ng/ml (n = 6). The data support the thesis that in differentiated thyroid carcinoma serum thyroglobulin levels are elevated when metastases develop after initial treatment. It is proposed that the measurement of thyroglobulin in the serum represents a simple and valuable adjunct in the posttreatment follow-up of patients with differentiated thyroid cancer.     

Microfluorometric demonstration of thyroglobulin antibodies with the defined antigen substrate spheres (DASS) system.]

Previous investigations showed that quantitative immunofluorescence using antigens covalently bound to agarose particles represents a reproducible and sensitive assay for antibodies in experimental antisera. In this paper data are presented which show that also thyroglobulin antibodies in patients sera can be demonstrated using the defined antigen substrate spheres (DASS) system with thyroglobulin as antigen. Sera of 45 patients with various thyroid diseases were investigated for the presence of thyroglobulin antibodies using passive haemagglutination and the quantitative immunofluorescence technique. Comparable results were obtained with both techniques. Advantages and disadvantages of both methods are discussed.     

Highly sensitive assays of autoantibodies to thyroglobulin and to thyroid peroxidase

These highly sensitive assays are based on the interaction between thyroid autoantibodies and 125I-labeled autoantigens. Serum samples are incubated with labeled thyroid peroxidase (TPO) or thyroglobulin (Tg) to allow the formation of antibody-labeled antigen complexes. The complexes are then precipitated by addition of solid-phase Protein A. In the presence of high concentrations of TPO antibody or Tg antibody, more than 50% of the respective labeled antigen was precipitated, whereas only 1-2% was precipitated in the absence of autoantibody. Interassay CVs were 3.2% and 5.7%, respectively, for the anti-TPO and anti-Tg assays. There was no cross-reactivity between Tg antibody and TPO antibody. Results correlated highly significantly with results from other assay systems based on antigen-coated cells or plastic supports, but the assays described here were considerably more sensitive. Scatchard analysis of the assay data provided information on the affinity and serum concentration of TPO autoantibodies (ka approximately 10(9) L/mol and concentrations up to 1 g/L) and Tg autoantibodies (ka approximately 4 x 10(10) L/mol and concentrations up to 1 g/L). Overall, these assays provide a sensitive, precise, and convenient system for measuring and investigating the properties of thyroid autoantibodies.     

Purification of human thyroid peroxidase using ion exchange liquid chromatography

We have utilized a one step ion-exchange (FPLC Mono Q) purification procedure for the isolation of human thyroid peroxidase (TPO). The purified TPO had the properties of a major microsomal antigen and inhibited the binding of human microsomal autoantibodies to thyroid microsomal membranes. The isolated TPO was free from thyroglobulin and showed, compared with crude microsomal proteins, a reduced background binding with control sera in enzyme-linked immunoassay (ELISA). SDS-gel electrophoresis of the isolated TPO detected one major band with an apparent molecular weight of 105 kD. The antigenicity of the protein was demonstrated by immunoblotting using sera from patients with autoimmune thyroiditis. These results demonstrate that FPLC Mono Q chromatography offers a rapid, quantitative and precise method for large scale purification of TPO with retained enzymatic and antigenic activity for use in ELISA and for further studies on the structure and function of this protein.     

Concordance of eight kits for antithyroid peroxidase autoantibodies determination.

Numerous methods are proposed to quantify antithyroid peroxidase autoantibodies. No standardization exists but most assays use the standard MRC 66/387 with a calibration factor. Costs of the tests vary between the different kits. We evaluated the concordance of eight peroxidase autoantibodies assay kits in two centres, using a panel of sera from 269 subjects: controls (n=100), patients with autoimmune thyroid disease (n=77; Graves' disease, Hashimoto's thyroiditis), patients with non-autoimmune thyroid disease (n=69; nodular goiter, differentiated thyroid carcinoma) and individual sera with thyroglobulin antibodies only (n=23). The concordance between the eight methods was high, ranging from 88.3% to 98.8% with the total panel of sera. The majority of assays demonstrated high diagnostic performance. We encountered some false-positive results at borderline positive levels, and the nonrecognition of some sera by competitive assays.     

Thyroglobulin: a specific serum marker for the management of thyroid carcinoma

Thyroglobulin measurements in tissue and serum play an integral role in the evaluation of patients who have thyroid cancer. Immunohistochemical detection of thyroglobulin in surgical specimens is useful in the differential diagnosis of tumors of unknown origin; however, the most important application of thyroglobulin measurement in clinical practice is in the postsurgical management of differentiated thyroid cancer. Serum thyroglobulin is a highly specific and sensitive tumor marker for detecting persistent or recurrent thyroid cancer and for monitoring clinical status. The reappearance of circulating thyroglobulin after total thyroid ablation is pathognomonic for the presence of tumor. The measurement of thyroglobulin in serum is challenging, however, and several analytical problems limit assay performance. Thyroglobulin autoantibody interference is a particularly significant concern that requires all thyroglobulin samples to be screened for their presence. No immunoassay is totally free from interference by thyroglobulin autoantibodies. Measurement of thyroglobulin mRNA to detect circulating tumor cells may help to overcome some of the limitations of current protein-detection methods; serum thyroglobulin will continue to remain the "gold standard." The complex functional features of thyroid carcinomas make sole reliance upon any one diagnostic technique, including thyroglobulin assessments, potentially misleading. Thyroglobulin measurements are a critical component of a multifaceted diagnostic approach to this disease.     

Distinction between epidermal antigens binding pemphigus vulgaris and pemphigus foliaceus autoantibodies.

Pemphigus vulgaris (PV) and pemphigus foliaceus (PF) are autoimmune blistering diseases in which antibodies develop to the cell surface of epidermal cells. In this study we sought to determine the antigenic specificity of antibodies in the sera of patients with PV and PF. Sera from 12 patients with PV were used to immunoprecipitate extracts of cultured human epidermal cells that were radiolabeled with 14C-amino acids. Immunoprecipitates were identified by SDS polyacrylamide gel electrophoresis (PAGE) and fluorography. All 12 PV sera precipitated a protein which, when reduced, displayed chains of 130,000 and 80,000 mol wt on SDS-PAGE. Electrophoresis under nonreducing conditions identified a 210,000-mol wt molecule, which was presumably formed by disulfide crosslinking of the 130,000 and 80,000-mol wt chains. Immunoprecipitates of epidermal cell extracts that were labeled with 14C-glucosamine indicated that the 130,000-mol wt chain. Seven of eight PF sera, which were run concurrently with the PV sera in this immunoprecipitation assay, did not precipitate this glycoprotein, nor did they specifically precipitate any protein. To determine if a specific molecule which reacted with antibodies in PF sera could be identified, we used immunoblot analysis of extracts of normal human epidermis. The proteins in these extracts were reduced, separated by SDS-PAGE, and electrophoretically transferred to nitrocellulose sheets or to 2-aminophenylthioether paper. Immunoperoxidase staining of the transferred proteins with PF sera indicated that four of eight PF sera contained antibodies that stained a protein band of 160,000 mol wt. Indirect immunofluorescence, using normal human skin as the substrate, indicated that IgG that was eluted from this protein band stained the epidermis in a cell surface pattern. PV sera did not specifically recognize any bands by immunoblot analysis. Immunoblots performed with PV antigen that was immunoprecipitated from cell culture extracts suggested that, once denatured for SDS-PAGE, PV antigen is no longer immunoreactive. Taken together, these data indicate that: autoantibodies contained in PV sera from various patients have a unique molecular specificity; autoantibodies from most PF sera have a specificity different from that of PV autoantibodies; and autoantibodies from various PF patients may not have identical antigenic specificities. These differences in antigenic specificity between PV and PF sera may account for the clinical and histologic differences between these diseases.     

The detection of a unique antigen associated with papillary thyroid carcinoma.

We produced antibodies against a thyroid papillary carcinoma homogenate (PCAb) and analyzed antigens recognized by this antibody using western blotting. Fifty-four thyroid tissue specimens and 6 control tissue specimens obtained from non-thyroid carcinoma (gastric tissue, colon and liver) were analyzed. Consequently, an antigen of 40 kDa in size was found in 16 of 16 (100%) of papillary thyroid carcinoma from primary lesions and in 2 of 2 (100%) papillary thyroid carcinoma from metastatic foci, whereas it was not detected in thyroid tissue samples from follicular carcinoma, anaplastic carcinoma, follicular adenoma, adenomatous goiter, Graves' disease and normal thyroid tissues. The reactivity of thyroglobulin antiserum (TgAb) to this 40 kDa antigen was tested by western blotting and showed that TgAb did not appear to recognize the 40 kDa antigen. Moreover PCAb, after treatment with Tg, still reacted with this 40 kDa antigen. Therefore, this 40 kDa antigen might be different from Tg. Furthermore, to inspect the structure of this antigen, the effect of some chemicals and enzymes such as 2-mercaptoethanol, sodium dodecyl sulfate, ethanol and protease on the reactivity of PCAb to the 40 kDa antigen were analyzed. The results of these experiments suggested that this 40 kDa antigen may have a peptide structure. To our knowledge, the finding reported here represents the first demonstration of the protein specifically present in papillary thyroid carcinoma. Further investigations should elucidate the characteristics of this antigen and may contribute to definitive diagnosis of thyroid carcinoma as well as improving the understanding of the mechanisms involved in developing the thyroid carcinoma.     

Insights into native epitopes of proliferating cell nuclear antigen using recombinant DNA protein products

A cDNA clone encoding full-length human proliferating cell nuclear antigen (PCNA) was used to generate a panel of in vitro translated labeled protein products with COOH-terminal deletions and to construct a set of fusion proteins with COOH- and NH2-terminal deletions. A rabbit antiserum raised against an NH2-terminal peptide, a well-characterized murine monoclonal antibody (mAb), and 14 human lupus sera with autoantibody to PCNA were analyzed for their reactivity with the constructs using both immunoprecipitation and immunoblotting techniques. The rabbit antiserum reacted in immunoprecipitation and immunoblotting with constructs containing the appropriate NH2-terminal sequence and mAb reacted with a sequence from the midregion of PCNA. These experimentally induced antibodies also reacted with 15-mer synthetic peptides in enzyme-linked immunosorbent assay (ELISA). In contrast, none of the lupus sera reacted with synthetic peptides in ELISA. 9 of the 14 lupus sera also failed to react in Western immunoblotting with any recombinant fusion protein, although they all immunoprecipitated in vitro translated full-length protein. Four of the nine had variable patterns of immunoprecipitation with shorter constructs. The remaining five lupus sera were able to immunoprecipitate translation products as well as Western blot recombinant fusion proteins. From analysis of the patterns of reactivity of human lupus sera, it was deduced that the apparent heterogeneity of human autoantibodies to PCNA could be explained by immune response to highly conformational epitopes. These observations demonstrate that there might be special features in "native" epitopes of intranuclear antigens that are recognized by autoantibodies, and that these special features of native epitopes might not be present in prepared antigen used for experimental immunization. These features may be related to protein folding or to association of the antigen with other intranuclear proteins or nucleic acids, as might occur with antigens that are components of subcellular particles.     

Serum thyroglobulin and its autoantibody following subtotal thyroid resection of Graves'' disease

Thyroid surgery leads to marked changes of the levels of serum thyroglobulin and its autoantibodies in the subsequent 3 postoperative weeks. Furthermore in Graves' disease progression of exophthalmos has sometimes been seen following thyroidectomy. Nineteen medically pretreated patients with Graves' disease and no signs of exophthalmos were studied systematically up to 6 months postoperatively. Nine patients had thyroglobulin antibodies. Mean values rose to 3.5 times pretreatment values within 2 months (P less than 0.001) followed by a gradual fall below pretreatment level after 6 months. None of the antibody negative patients reverted to positive or vice versa. Serum thyroglobulin (n = 10) was elevated preoperatively (mean 309 micrograms/l, SD 251), their values being normalized within 1-2 months (mean 19.4 micrograms/l, SD 7.3). The preoperative serum thyroglobulin correlated to the weight of the removed thyroid tissue (r = 0.87, P less than 0.01). Three patients showed elevated thyroid stimulating hormone after 1 month. Of these, two developed myxoedema, the third remained euthyroid with persistently elevated serum thyroglobulin. None showed recurrence or developed exophthalmos within the period of observation. In spite of rising levels of thyroglobulin antibodies in all patients with antibodies none developed exophthalmos and only one patient with thyroglobulin antibodies had clinical myxoedema.     

Autoantibodies in complex regional pain syndrome bind to a differentiation-dependent neuronal surface autoantigen

Complex regional pain syndrome, which is characterised by pain and trophic disturbances, develops frequently after peripheral limb trauma. There is an increasing evidence of an involvement of the immune system in CRPS, and recently we showed that CRPS patients have autoantibodies against nervous system structures [6]. Therefore we tested the sera of CRPS patients, neuropathy patients and healthy volunteers for surface-binding autoantibodies to primary cultures of autonomic neurons and differentiated neuroblastoma cell lines using flow cytometry. Thirteen of 30 CRPS patients, but none of 30 healthy controls and only one of the 20 neuropathy sera had specific surface binding to autonomic neurons (p < 0.001). The majority of the sera reacted with both sympathetic and myenteric plexus neurons. Interestingly, 6/30 CRPS sera showed binding to undifferentiated SH-SY5Y neuroblastoma cells. However, differentiation of SH-SY5Y into a cholinergic phenotype induced a surface antigen, which is recognised by 60% of CRPS sera (18/30), but not by controls (p < 0.001). Our data show that about 30–40% of CRPS patients have surface-binding autoantibodies against an inducible autonomic nervous system autoantigen. These data support an autoimmune hypothesis in CRPS patients. Further studies must elucidate origin and function of these autoantibodies in CRPS.     

Systemic lupus erythematosus patients with central nervous system involvement show autoantibodies to a 50-kD neuronal membrane protein

An antibody was detected in the sera from patients with systemic lupus erythematosus (SLE) and central nervous system (CNS) involvement that reacted with a 50-kD antigen in the plasma membrane of brain synaptic terminals. The 50-kD antigen was solubilized with Triton X-100 from preparations enriched with synaptic plasma membranes, and was partially purified by molecular sieve filtration column chromatography. The sera of 19 of 20 CNS-SLE patients showed strong to moderate immunoreactivity with the 50-kD protein in Western blots. Immunoreactivity with the 50-kD protein was also detected in the cerebrospinal fluid of CNS-SLE patients. Control sera from healthy individuals did not react with the 50-kD protein. Low to background reactivity was detected in 35% of a group of SLE patients without CNS manifestations, and in 3% of patients displaying other connective tissue diseases. A total of 100 individuals were tested in this study. Purified autoantibodies to the 50-kD protein from CNS-SLE patients were used for immunofluorescent labeling of neuroblastoma cells. The immunofluorescent staining revealed a distinct macular distribution pattern on the surface of the cell membrane. Taken together, the data suggest that the 50-kD protein may be an important target for autoantibodies, preponderantly found in CNS-SLE patients, and that the antigen may play a role in the pathogenesis of some neurological manifestations in SLE.     

基于免疫金银染色的蛋白质与抗原分子微阵列技术

目的　研制适于自身抗体检测的蛋白质与抗原分子微阵列。方法　利用氧化型琼脂糖凝胶膜结合戊二醛分子衍生的活性表面 ,制作蛋白质与抗原分子微阵列 ,建立了检测血清自身抗体的胶体金免疫金银染色方法 (IGSS)。结果　在未经选择的风湿病患者中 (包括SLE ,RA和MCTD) ,检出多种自身抗体阳性 ,其中又以子宫内膜抗体 (EmAb)、抗ssDNA抗体、抗TG抗体、抗TPO抗体和RF等的检出频率较高 ;但心磷脂抗体 (ACA)、抗LSP和抗精子抗体 (AsAb)均为阴性。经初步方法学对比考证 ,实验结果与ELISA有较好的一致性 (达 90 % )。结论　本法具有方法敏感 ,结果直观和实验条件易于控制等特点 ,为发展可视化阵列芯片提供了一种很好的模式。