Dot-IGSS法检测SLE患者血清中抗肾小球基底膜及肾小管抗体的研究

方法：本实验采用斑点免疫金银染色（Ｄｏｔ－ＩＧＳＳ）法对５０例ＳＬＥ患者血清中的肾小球基底膜抗体（ＧＢＭ－Ａｂ）及肾小管抗体（Ｔｕｂ－Ａｂ）进行检测。结果：ＧＢＭ－Ａｂ阳性率达６２％，Ｔｕｂ－Ａｂ阳性率达７６％。与患者尿蛋白检查（阳性率为４８％）比较，上述自身抗体检出率较高。此外，ＧＢＭ－Ａｂ或Ｔｕｂ－Ａｂ检测阳性患者与肾活检诊为狼疮性肾炎（ＬＮ）相符率达１００％。结论：ＧＢＭ－Ａｂ或Ｔｕｂ－Ａｂ的检出可为临床早期诊断ＬＮ提供可靠的证据     

Dot—IGSS法检测SLE患者血清中抗肾小球基底膜及肾小管抗？…

方法；本实验采用斑点免疫金银染色（Ｄｏｔ－ＩＧＳＳ）法对５０例ＳＬＥ患者血清中的肾小球基底膜抗体（ＧＢＭ－Ａｂ）及肾小管抗体（Ｔｕｂ－Ａｂ）进行检测。结果：ＧＢＭ－Ａｂ阳性率达６２％，Ｔｕｂ－Ａｂ阳性率达７６％。与患者尿蛋白检查比较，上述自身抗体检出率较高。此外，ＧＢＭ－Ａｂ或Ｔｕｂ－Ａｂ检测阳性患者与肾活检诊为狼疮性肾炎（ＬＮ）相符率达１００％。结论：ＧＢＭ－Ａｂ或Ｔｕｂ－Ａｂ的检出可为临床早期     

1984-1995年12年间西安地区孕妇TORCH特异性抗体检测结果分析

本文报告自１９８４年１月至１９９５年５月间西安地区孕妇ＴＯＲＣＨ特异性抗体检测结果分析。本室用微量间接血凝法及ＥＬＩＳＡ法对孕妇血清分别检测弓形体抗体（ＴＰ－Ａｂ）３１４４例、抗风疹病毒ＩｇＭ（ＲＶ－ＩｇＭ）９２０例、抗巨细胞病毒ＩｇＭ（ＣＭＶ－ＩｇＭ）１４５４例、抗单纯疱疹病毒Ⅱ型ＩｇＭ（ＨＳＶⅡ－ＩｇＭ）９５９例，其结果阳性率分别为３．６６％０．１１％、１１．２８％、３．０２％。其中，早孕和中晚孕孕妇间的各项特异性抗体阳性率均无显著性差异（Ｐ＞０．０５）；有无上感病史孕妇间的ＴＰ－Ａｂ、ＣＭＶ－ＩｇＭ、ＨＳＶⅡ－ＩｇＭ阳性率均无显著性差异（Ｐ＞０．０５）；不同职业孕妇的ＴＰ－Ａｂ阳性率亦无显著性差异（Ｐ＞０．０５）。从而得知，对所有的孕妇在孕期进行ＴＯＲＣＨ感染检测，对于作好优生工作有重要的意义。     

自身免疫病患者抗精浆免疫抑制物抗体的测定

用ＥＬＩＳＡ法测定血清中抗精浆免疫抑制物抗体（ＳＰＩＭ－Ａｂ），发现系统性红斑狼疮、类风湿性关节炎、混合性结缔组织病、重症肌无力和甲状腺疾病患者血清ＳＰＩＭ－Ａｂ水平与检出率均显著高于对照组（Ｐ＜０．０１）；抗核抗体、抗ｄｓ－ＤＮＡ抗体、抗Ｓｍ、抗ＲＮＰ抗体和类风湿因子与ＳＰＩＭ－Ａｂ无相关性，而抗甲状腺球蛋白抗体（Ａ－ＴＧ）和抗甲状腺微粒体抗体（Ａ－ＴＭ）阳性患者ＳＰＩＭ－Ａｂ水平均显著高于Ａ－ＴＧ、Ａ－ＴＭ阴性者（Ｐ＜０．０１）。     

Dot—IGSS和Dot—ELISA法检测GBM—Ab和Tub—Ab的比较研究

本实验应用斑点免疫金银染色及斑点酶联免疫吸附试验对５０例ＳＬＥ患者体内的肾小球基底膜抗体及肾小管抗体进行了检测比较。结果表明：Ｄｏｔ－ＩＧＳＳ法检测ＧＢＭ－Ａｂ和Ｔｕｂ－Ａｂ的阳性率分别是６２％和７６％，Ｄｏｔ－ＥＬＩＳＡ法检测ＧＢＭ－Ａｎ和Ｔｕｂ－Ａｂ的阳性率分别是６８％和７８％。     

多抗F（ab‘）2—ELISA测定人血清抗TG—Ab2及意义

为了克服单抗测定甲状腺球蛋白独特型抗体的局限性，便于常规检测，我们建立了兔抗ＴＧ多克隆抗体的Ｆ（ａｂ）２－ＥＬＩＳＡ测定人血清中的ＴＧ－Ａｂ２。结果其抗ＴＧ－Ａｂ２阳性率；甲亢为９．９％，桥本氏甲状腺炎为４４．４％，甲状腺瘤为１６．７％，ＳＬＥ为９．５％，类风湿性关节炎为０％，正常人为０％。     

眼肌结合抗体的测定及其致免疫损伤机制的研究

用牛和猪眼肌抗原建立了检测眼肌结合抗体（ＥＭｂ Ａｂ ） 的ＥＬＩＳＡ法。证实甲亢患者血清中存在ＥＭｂ Ａｂ。通过ＥＭｂ Ａｂ 特异免疫复合物、补体复合物解离活性抗体依赖的细胞毒或补体介导的细胞毒（ＡＤＣＣ或ＣＭＡＣ） 测定, 以及ＥＭｂ Ａｂ 与眼肌细胞抗原结合的实验观察, 表明此种ＥＭｂ Ａｂ 可能在与已受某种因素损伤后的眼肌结合时, 通过ＡＤＣＣ反应加剧眼肌损伤。     

多抗F（ab＇）_2－ELISA测定人血清抗TG－Ab_2及意义

为了克服单抗测定甲状腺球蛋白独特型抗体（抗ＴＧ－Ａｂ＿２）的局限性，便于常规检测，我们建立了兔抗ＴＧ多克隆抗体的Ｆ（ａｂ＇）＿２－ＥＬＩＳＡ测定人血清中的抗ＴＧ－Ａｂ＿２．结果其抗ＴＧ－Ａｂ＿２阳性率：甲亢（Ｇｒａｖｅ＇３）为９．９％（７／７１），桥本氏甲状腺炎为４４．４％（４／９），甲状腺瘤为１６．７％（３／１８），ＳＬＥ为９．５％（４／４２），类风湿性关节炎为０％（０／３２），正常人为０％（０／３５）。     

西安地区死胎、流产的孕妇微小病毒B19抗体检测结果分析

用ＥＬＩＳＡ法对孕早期有死胎、流产的５７例孕妇（实验组）与３５例正常孕妇（对照组）血清检测Ｂ１９抗体（ＩｇＭ、ＩｇＧ）其中实验组Ｂ１９－ＩｇＭ阳性率（１２．２８％）与正常组Ｂ１９－ＩｇＭ阳性率比较有显著性差异（ｐ＜０．０５），实验组Ｂ１９－ＩｇＧ阳性率（３３．３３％）与正常组Ｂ１９－ＩｇＧ阳性率（４０％）无统计学差异（Ｐ＞０．０５）。     

SLE患者血清抗眼肌抗体的检测

ＳＬＥ患者血清抗眼肌抗体的检测马东生，王艾丽，武建国南京军区南京总医院南京２１０００２抗眼肌抗体（ＥＭ－ａｈ）是一组对眼肌细胞膜多种膜蛋白抗原起反应的自身抗体，对眼肌细胞具有细胞毒作用［１］。为探讨ＳＬＥ眼病与ＥＭ－ａｈ的关系，我们用间接ＥＬＩＳＡ法...     

Comparative study on IgG and IgA antibodies against human thyroid and eye-muscle antigens in Graves' ophthalmopathy.

Circulating IgG and IgA anti-thyroid and anti-eye muscle antibodies were investigated in 87 patients with Graves' disease (60 cases with ophthalmopathy). The ELISA method was used. Both IgG and IgA antibodies were demonstrated against human thyroid and eye-muscle membrane or cytosol antigens. Anti-eye-muscle antibodies of the IgA type were observed more frequently than those of the IgG type (25 cases vs. 18 were demonstrated with membrane antigens and 37 cases vs. 23 with cytosol antigens). The respective distributions for thyroid antigens the cytosol fraction were 55 cases vs. 13 and 18 cases vs. 36. A significant difference was observed in the anti-thyroid IgG levels and the anti-eye-muscle membrane or cytosol levels between the patients with Graves' disease and those in control group (P less than 0.001). The difference in the IgA antibody to thyroid and eye-muscle antigens was significant between the patients with and without ophthalmopathy (P less than 0.002). The strong correlation between the levels of IgA antibodies to thyroid and those to the eye-muscle cytosol fractions might be connected with the theory of the common aetiology of the thyroid and eye diseases in Graves' ophthalmopathy (P less than 0.001). Circulating IgA anti-human thyroid and eye-muscle antibodies seemed to have a diagnostic relevance in the development of ophthalmopathy in Graves' ophthalmopathy.     

Frequent anti-V-region immune response to mouse B72.3 monoclonal antibody

The immune response of 56 colorectal cancer patients to a single infusion of 1 mg of radiolabeled (¹¹¹In) mouse B72.3-GYK-DTPA immunoconjugate was examined using a double-antigen radiometric assay system. The incidence of antibody response was 48% to polyclonal mouse IgG, 71% to mouse B72.3, and 62% to chimeric B72.3. Twelve patients (23%) had an antibody response to B72.3 V region in the absence of binding to polyclonal mouse IgG. An antiidiotype response was demonstrated in sera from 36% of 25 patients examined and correlated well with chimeric B72.3-GYK-DTPA immunoconjugate binding (r=0.72), moderately well with mouse B72.3 binding (r=0.56), and not at all with polyclonal mouse IgG binding (r=0.28). The peak antibody response occurred most frequently 2 weeks postinfusion, although a delayed peak response to chimeric B72.2 occurred in 29% of patients. This study suggests that mouse B72.3 causes an immune response in the majority of patients and that antibody response to the V region is common. Understanding the physiological significance of these antibody responses will require correlation with the kinetics and tumor localization of repeat infusions of such immunoconjugates.     

Inefficient Binding of IgM Immune Complexes to Erythrocyte C3b–C4b Receptors (CR1) and Weak Incorporation of C3b–iC3b into the Complexes

The binding of soluble complement-reacted IgM immune complexes (IC) to erythrocyte (E) C3b–C4b receptors (CRI) and the incorporation of C3b–iC3b into solid phase IgM-IC was investigated. The optimal binding of liquid phase IgM-IC to E-CRI was obtained with IC formed at moderate antibody excess, but the binding was low (2–3%) when compared to the binding of the corresponding IgG-IC (50–60%). Solid phase IC were prepared by coming microwells with heat-aggregated bovine serum albumin (BSA) followed by incubation with rabbit IgM anti-BSA antibody. The IC were reacted with human serum at 37°C. The binding of C3b–iC3b was determined by use of biotinylated F(ab')₂ antibodies to C3b-C3c and avidin-coupled alkaline phosphatase. The incorporation of C3b–iC3b into solid-phase IgM-IC increased when increasing amounts of IgM antibody were reacted with the antigen. The binding reaction was slow, reaching a maximum after about 2 h at 37°C. The binding of C3b–iC3b to the IgM-IC was remarkably inefficient when compared to the incorporation into IgG-IC reacted with the same amounts of BSA-precipitating antibody.     

Anti-RNA polymerase I antibodies in the sera of MRL lupus mice at the initial stages of disease are directed primarily against phosphorylation-dependent epitopes.

Anti-RNA polymerase I (RPI) antibodies in the sera of MRL/Mp-lpr/lpr and MRL/Mp(-)+/+ mice, which develop an autoimmune disease similar to human systemic lupus erythematosus, were screened for reactivity with purified RPI or RPI which had been dephosphorylated. In every case (n = 10), dephosphorylation of RPI resulted in a significant decrease (33-95%) in antibody binding. The anti-RPI antibodies in the sera of the same mice approximately 6 weeks later also reacted better with untreated as compared to dephosphorylated RPI but, in every case, the decrease in antibody (0-30%) caused by dephosphorylation was substantially diminished. That the proportion of anti-RPI antibodies in the sera of MRL mice decreased with progression of lupus-like disease was confirmed by closely monitoring the antibodies over the course of disease. Anti-RPI antibodies produced at the earliest stages appeared to be directed almost exclusively against phosphorylation-dependent determinants since dephosphorylation of RPI essentially abolished antibody binding. Subsequently, the percentage of the total anti-RPI antibodies in the sera of these mice directed towards phosphorylation-independent epitopes increased linearly with time. The importance of phosphorylation-dependent epitopes on RPI for the development of the anti-RPI autoimmune response was supported by the observation that treatment of mice with alkaline phosphatase partially attenuated anti-RPI antibody production.     

IgE antibody to cat allergens in an allergic population.

Clinical cat sensitivity and IgE antibody to cat allergens were studied in 200 consecutive patients who presented themselves for an allergy evaluation. Fifty-nine percent of the patients had no evidence of allergy to cats (group 1). Twenty-three percent of the patients had positive histories of allergy to cats and a positive prick test to cat epithelial extracts (group 2). The remaining 18% had either positive prick tests or positive histories alone (groups 3 and 4). The group of patients with strong evidence of allergy to cats (group 2) was on an average younger and had a higher incidence of asthma than did group 1 patients. Significant binding of IgE antibody to various insolubilized allergen preparations was defined as that binding which exceeded the 95% confidence interval for group 1 patients. Significant binding of IgE antibody to insolubilized cat allergen 1 was present in 65%, 15%, and 10% of groups 2, 3, and 4 patients, respectively. Significant binding of IgE antibody to insolubilized cat pelt extract was present in 28%, 11%, and 0% of group 2, 3, and 4 patients, respectively. Data for IgE antibody binding to insolubilized cat serum and cat albumin revealed no significant difference between the groups. It is concluded that cat allergen 1 is an important allergen in patients with strong evidence of allergy to cats.     

Immunoaffinity beads for selective removal of cholesterol from human plasma

Anti-low density lipoprotein antibody (anti-LDL antibody) attached poly(2-hydroxyethyl methacrylate-methacryloylamidophenylalanine) (poly(HEMA-MAPA)) beads were prepared for selective removal of cholesterol from hypercholesterolemic human plasma. Poly(HEMA-MAPA) beads were produced by a modified suspension polymerization and then characterized by swelling tests and SEM. Blood-compatibility tests were also investigated. The water swelling ratio of the poly(HEMA-MAPA) beads increased significantly (68%) compared with pHEMA (55%). All the clotting times increased when compared with poly(HEMA) beads. Loss of platelets and leukocytes was very low. The maximum anti-LDL antibody attachment was achieved at pH 7.0. Attachment of anti-LDL antibody was 29.6 mg/g. There was a very low non-specific cholesterol binding onto the poly(HEMA-MAPA) beads, about 0.74 mg/g. Anti-LDL antibody attached beads adsorbed in the range of 13.3-16.0 mg cholesterol/g from hypercholesterolemic human plasma. Up to 92% of the adsorbed LDL was desorbed. The binding-elution cycle was repeated 10 times using the same beads. There was no significant loss of binding capacity.     

Immunoaffinity beads for selective removal of cholesterol from human plasma

Anti-low density lipoprotein antibody (anti-LDL antibody) attached poly(2-hydroxyethyl methacrylate-methacryloylamidophenylalanine) (poly(HEMA-MAPA)) beads were prepared for selective removal of cholesterol from hypercholesterolemic human plasma. Poly(HEMA-MAPA) beads were produced by a modified suspension polymerization and then characterized by swelling tests and SEM. Blood-compatibility tests were also investigated. The water swelling ratio of the poly(HEMA-MAPA) beads increased significantly (68%) compared with pHEMA (55%). All the clotting times increased when compared with poly(HEMA) beads. Loss of platelets and leukocytes was very low. The maximum anti-LDL antibody attachment was achieved at pH 7.0. Attachment of anti-LDL antibody was 29.6 mg/g. There was a very low non-specific cholesterol binding onto the poly(HEMA-MAPA) beads, about 0.74 mg/g. Anti-LDL antibody attached beads adsorbed in the range of 13.3-16.0 mg cholesterol/g from hypercholesterolemic human plasma. Up to 92% of the adsorbed LDL was desorbed. The binding-elution cycle was repeated 10 times using the same beads. There was no significant loss of binding capacity.     

Factors Involved in Macrophage: Immune Complex Binding

The interaction of the immune complex (IC) composed of DNA and monoclonal anti-DNA antibody with thioglycollate-stimulated mouse peritoneal macrophages was investigated. The immune complex: macrophage interaction was shown to be highly time and temperature dependent; at 37°C it proceeds faster than at 0°C, although there is higher overall binding of IC to macrophages at 0°C. The maximum bound IC detected was at a DNA/antibody ratio of 6.2ng/ml to 7.3μg/ml. Higher densities of either DNA or antibody inhibited IC: phagocyte interaction.

Binding of the IC to macrophages is through cell surface Fc receptors and is enhanced in the presence of 40 mg/ml albumin. Fresh human and mouse sera at the concentration of 10 percent, inhibited the IC binding to mouse peritoneal macrophages. Macrophage receptors for IC are not saturated even after 60 minutes. Addition of either chloroquine or cytochalasin B, resulted in increased binding of IC to macrophages.     

Development of an immunobiosensor assay for the beta-adrenergic compound zilpaterol

A surface plasmon resonance biosensor method was developed to measure zilpaterol residues in sheep urine. A CM-5 sensor chip previously reacted with ethylenediamine to produce an aminoethyl group was coupled with 4-carboxybutyl zilpaterol activated using EDC/NHS. Five polyclonal and four monoclonal antibodies were screened for their suitability to detect low levels of zilpaterol using the biosensor technology. Total binding was greater for polyclonal than monoclonal antibodies, but a less diluted antibody solution was required for polyclonal antibodies. A fixed antibody concentration and various concentrations of zilpaterol were injected to obtain a standard curve for each antibody to allow for B₀ and IC₅₀ determination. The stability of the assay was assessed by the consistency of B₀ in repeated experiments extending at least six hours. A measure of non-specific binding allowed the assessment of the specificity of the antibody-immobilized ligand interaction. The effect of varying concentrations of urine on B₀ and IC₅₀ was evaluated to assess the degree of “matrix” effect that would be present in an assay. Based on these criteria the most promising antibody (2E10, a monoclonal antibody) was selected for further evaluation. This antibody had good sensitivity with IC₅₀=4.47±0.41 ng/ml (n=11) in buffer). Both intra- and inter-assay variation studies showed excellent recovery and reproducibility for concentrations between 2 ng/ml and 8 ng/ml. A comparison of the biosensor method with a previously developed ELISA demonstrated that both methods give equivalent results (slope of the correlation plot = 1.02) with a high correlation (r²=0.91) between them.     

Determination of avidity of anti-viral antibodies at 50% binding of antibody.

Binding studies with tobacco mosaic virus and specific IgG and Fab' fragments were done under conditions of 50% binding of antibody. When the log of the concentration of antigen sites required to bind 50% of the available antibody was plotted against the log of bound sites, a line was obtained which had a slope varying between 0 and 1. Instead of using the slope of this line as an index of avidity of the antiserum, it is suggested that the calculation K0 values over a range of reactant concentrations provides a better criterion for comparing the avidity of antiviral antibodies. The results of conventional binding tests were analyzed by means of Scatchard plots and were found to agree very closely with the data obtained at 50% binding of antibody.