抗细胞膜DNA抗体的测定及其在系统性红斑狼疮诊断中的意义

目的建立抗细胞膜DNA(mDNA)抗体测定的方法,研究其在系统性红斑狼疮(SLE)诊断中的敏感性和特异性,以及与SLE临床特点及免疫学异常的关系.方法检测SLE患者、疾病对照组和正常对照组血清中的抗mDNA抗体,并分析SLE患者的各种临床表现及实验室指标与抗mDNA抗体的关系.同时研究细胞膜DNA分子在不同细胞表面的表达,用DNA酶、RNA酶及胰酶鉴定抗原性质.结果抗mDNA抗体在SLE中检测敏感性73.3%(152/207),特异性96.4%,疾病对照组阳性率5.4%(9/167),82名正常对照组均为阴性,抗mDNA抗体阳性率在SLE组明显高于疾病对照组和正常对照组(P＜0.01).抗mDNA抗体在其他自身抗体阴性的SLE患者中有较高的检出率,在抗双链DNA(dsDNA)抗体、抗Sm抗体、快速狼疮因子(DNP)、抗组蛋白抗体(AHA)、抗核小体抗体(AnuA)阴性的SLE患者中抗mDNA抗体的阳性率分别是73.8%、62.7%、65.3%、57.8%和51.6%.该抗体阳性组SLE患者皮疹,脱发,关节痛,白细胞和C3、C4减低及IgG、IgA、IgM升高较为常见,但与SLE患者病情活动指数无关.本文还证实细胞膜DNA在人B细胞、T细胞上均有表达,以Raji细胞株表达较好.用DNA酶预处理的细胞涂片再行检测后膜荧光图形消失,而用RNA酶、胰酶预处理后并不消失,证实其为膜DNA抗原.结论抗mDNA抗体是一种诊断敏感性高、特异性强的SLE血清学指标之一,尤其对抗dsDNA、抗Sm、抗DNP、AHA、AnuA阴性的SLE的诊断有参考意义.     

抗细胞膜DNA自身抗体快速检测系统性红斑狼疮

目的 探讨抗细胞膜DNA自身抗体（mDNA)在系统性红斑狼疮患者的表达以及这种特异抗体与抗dsDNA抗体的不同。方法 用间接免疫荧光法观察培养的HL60细胞的特异性细胞膜的荧光图形。结果 抗细胞膜DNA自身抗体在SLE患者90例中有81例阳性（阳性率90％）,而在类风湿关节炎（RA）患者35例,强直性脊柱炎（AS）患者31例和健康献血员42名中均为阴性,在干燥综合征（SS）患者20例中有1例阳性（阳性率5％）,抗细胞膜DNA自身抗体对SLE诊断的特异性为98．8％。结论 抗细胞膜DNA自身抗体是一种诊断。SLE的新的标记抗体,其敏感性高（90．0％）,特异性强（98．8％）,较抗dsDNA抗体、抗Sm抗体检测更先进。     

抗细胞膜DNA抗体检测方法的建立及对系统性红斑狼疮诊断价值的临床应用研究

目的 应用间接免疫荧光法(IIF)检测抗细胞膜DNA(cmDNA)抗体,探讨抗cmDNA抗体对系统性红斑狼疮(SLE)的诊断价值.比较以人B淋巴细胞株Raji、人早幼粒白血病细胞株HL60为底物,抗cmDNA抗体在SLE中的检测效果.方法 选取306例SLE患者,192例非SLE疾病对照组,包括脊柱关节病(SPA) 52例,类风湿关节炎(RA) 70例,原发干燥综合征(pSS) 30例,其他结缔组织病(CTD)40例.50名健康志愿者作为对照组.采用IIF法观察培养的人B淋巴细胞株Raji、人早幼粒白血病细胞株HL60的细胞膜的荧光图形;用IIF法检测抗核抗体、抗双链DNA(dsDNA)抗体;联合应用免疫双扩散法和蛋白印迹法检测抗Sm抗体,酶联免疫吸附试验(ELISA)法测定抗核小体抗体(AnuA).配对资料比较用McNemarX2检验,相关性分析采用Spearman相关及Logistic回归分析.结果以Raji及HL60 2种细胞株为底物检测SLE患者抗cmDNA抗体,敏感性分别为72.5％、76.1％,特异性分别为91.7％、86.8％,差异均无统计学意义(P＞0.05).抗cmDNA抗体的敏感性明显高于抗Sm抗体及抗dsDNA抗体,差异有统计学意义(P＜0.01);特异性与抗dsDNA抗体相似(P＞0.05),日低于抗Sm抗体及AnuA(P＜0.01).抗cmDNA抗体分别与抗dsDNA抗体、抗Sm抗体及AnuA联合检测在SLE诊断中的敏感性均明显高于上述自身抗体单独检测,差异有统计学意义(P＜0.05).抗cmDNA抗体与SLE患者的黏膜溃疡之间有相关性(OR=2.343,P=0.029).抗cmDNA抗体与红细胞沉降率(ESR)有相关性(OR=1.031,P=0.012).抗cmDNA抗体与SLE疾病活动指数(SLEDAI)评分无相关性(r=0.070,P=0.600).本文研究还证实Raji细胞株较HL60细胞株更易复苏、荧光图形亮度更强,表达效果更好.结论 抗cmDNA抗体对SLE诊断的敏感性高,特异性强,可能成为SLE诊断的相对特异性抗体之一.抗cmDNA抗体与抗dsDNA抗体、抗Sm抗体及AnuA联合检测可提高对SLE诊断的敏感性.选择Raji细胞株为底物检测抗cmDNA抗体较HL60细胞株更有优势.     

自身抗体联合检测在系统性红斑狼疮诊断中的意义

目的研究抗细胞膜DNA(cmDNA)抗体、抗核小体抗体(AnuA)、抗脱氧核糖核蛋白(DNP)抗体及抗双链DNA(dsDNA)抗体等特异性抗体在系统性红斑狼疮(SLE)诊断中的意义,并与抗核抗体(ANA)的敏感性和特异性进行比较。了解特异性自身抗体联合检测在SLE的诊断及临床应用中的意义。方法测定了125例SLE及118例疾病对照组(包括原发性干燥综合征、多发肌炎、系统性硬化症、未分化结缔组织病、类风湿关节炎、骨关节炎、强直性脊柱炎及银屑病关节炎)患者血清中的自身抗体。利用间接免疫荧光法测定抗cmDNA抗体和ANA,酶联免疫吸附试验(ELISA)测定AnuA、乳凝法检测抗DNP抗体、金标法测定抗dsDNA抗体。结果AnuA、抗cmDNA抗体、抗DNP抗体、抗dsDNA抗体和ANA在SLE患者中的阳性率分别为68%、38.4%、51.2%、49.6%和95.2%,均明显高于疾病对照组(3.4%、4.2%、1.7%、0.8%和25.4%),差异有统计学意义(P<0.001)。ANA与AnuA的敏感性显著高于其他三种抗体,差异有统计学意义(P<0.05);AnuA、抗cmDNA抗体、抗DNP抗体、抗dsDNA抗体和ANA的特异性分别为95.8%、96.6%、98.3%、99.2%和74.6%;AnuA在抗DNP抗体、抗dsDNA抗体阴性的SLE中的阳性率明显高于其他抗体(P<0.05);自身抗体的联合检测可提高SLE诊断的敏感性,但特异性无明显改变。结论抗cmDNA抗     

系统性红斑狼疮患者血清5种抗细胞核及细胞浆抗体检测及其意义

采用间接免疫荧光法检测60例系统性红斑狼疮(SLE)、30例其他结缔组织病及30例正常对照者血清中的抗核抗体(ANA)水平,用酶联免疫印记法检测抗Sm抗体、抗dsDNA抗体、抗核小体抗体(AnuA)、抗组蛋白抗体(AHA)和抗核糖体P蛋白抗体(APPA).发现SLE组抗Sm抗体、抗dsDNA抗体、AnuA、AHA及APPA的阳性率明显高于病例对照组及正常对照组;SLE组ANA的阳性率为98%,且滴度83.5%≥1:320,更有38.3%≥1:10 000.认为联合检测多种抗体可以避免因单项检测出现的漏诊,提高了对SLE诊断的敏感性和诊断效率.     

SmD1抗体检测在系统性红斑狼疮诊断中的意义

目的 研究抗SmD1抗体及其他特异性抗体在系统性红斑狼疮（SLE）诊断中的意义。方法 测定了44例SLE及136例非SEE（包括干燥综合征、未分化结缔组织病、强直性脊柱炎、类风湿关节炎）患者血清中的自身抗体。利用免疫印迹法测定抗SmD1抗体、抗核小体抗体（ANuA）和抗SSA60000抗体，间接免疫荧光法测定抗核抗体（ANA）和抗双链DNA（dsDNA）抗体．免疫斑点法检测抗Sm抗体。结果 抗SmD1抗体在SLE患者中的阳性率为47.7％，高于抗Sm抗体阳性率18．2％，差异有统计学意义（P〈0．05），而且其抗SmD1抗体的特异性达97.1％。结论 抗SmD1抗体在狼疮的诊断中有很高的特异性，敏感性也高于抗Sm抗体。5种特异性抗体在SLE诊断中有明显的互补作用，联合检测可明显提高其对SLE诊断的敏感性。     

系统性红斑狼疮血清抗核小体抗体水平及意义的探讨

目的 研究抗核小体抗体(AnuA)在系统性红斑狼疮(systemic lupus erythematosus，SLE)患者血清中的水平及其相关影响因素，探讨AnuA在SLE诊治中的作用和意义。方法 采用酶联免疫吸附法(EUSA)测定120例初诊SLE患者、55例其他风湿性疾病和30名健康对照血清中AnuA水平。同时记录各种临床表现，检测并分析其治疗前的其他自身抗体和实验室指标。结果 自身抗体在SLE及其他风湿病对照组的阳性率分别为AnuA56％和7％，抗dsDNA抗体35％和1％，抗Sm抗体24％和0。AnuA与SLE患者性别、年龄、病程无相关性，对sLE的诊断敏感性和特异性分别为55．8％、9513％，对狼疮肾炎(1upus nephritis，LN)的诊断敏感性和特异性分别为77．％、64．5%。AnuA与肝脏损害和疾病活动呈线性相关(t=3．152，2．171，P＜O．05)。AnuA分别与抗dsDNA、抗Sm联合检测对SLE诊断敏感性提高27％、40％（X^2值分别为38．930、18．161，P＜O．01)。结论AnuA在SLE血清中水平显著增高，AnuA测定是SLE诊断和治疗监测中有价值的新的实验室检测指标之一，与抗dsDNA抗体联合检测可提高诊断SLE、LN的敏感性和特异性。     

抗核小体抗体诊断系统性红斑狼疮的价值

280份血清,取自健康体检者50例，类风湿关节炎（RA）74例，系统性红斑狼疮（SLE）106例．其他结缔组织病50例．分别检测其抗核抗体（ANA），抗双链DNA（dsDNA），抗史密斯（Sm），抗组蛋白（His）及抗核小体抗体（AnuA）抗体。结果5种抗体诊断SLE的敏感性和特异性为；ANA96．2%、31％，AnuA80．1％、93％，dsDNA56．6%、92％，Sm49．7％、90％，His54．7％、85%。ANA、AnuA敏感性明显高于其他3种抗体（P〈0．001）．AnuA、dsDNA、HiS、Sm的特异性明显高于ANA（P〈0．001）。AnuA是SLE的又一种特异性抗体，其与ANA、dsDNA抗体及Sm抗体、His抗体联合检测对SLE的诊断有重要意义。     

抗核小体抗体对系统性红斑狼疮的诊断价值

系统性红斑狼疮 (systemiclupuserythematosus,SLE)是一种常见的自身免疫疾病 ,它的主要特征是体内产生抗多种核抗原的自身抗体。包括DNA、组蛋白和核小体等。目前临床实验室诊断SLE主要采用抗核抗体 (ANA)、抗dsDNA抗体、抗可提取的核抗原 (ENA)抗体的检测。ANA是一种非特异性自身抗体。抗ENA谱中的抗Sm抗体虽然对SLE有很高的特异性 ,但敏感性较差 ,只有 30 %的阳性率 ;抗dsDNA抗体虽然对SLE具有较高的特异性和敏感性 ,但也有相当部分的SLE患者抗dsDNA抗体阴性。近来抗核小体抗体的检测据报道其特异性、敏感性均与抗dsDN…     

儿童系统性红斑狼疮患者血清抗核小体抗体检测

目的探讨抗核小体抗体(anti-nucleosome antibodies,AnuA)对儿童系统性红斑狼疮(systemic lupus erythematosus,SLE)诊断的意义,并与成人SLE进行比较。方法选取87例儿童SLE患者,30例非SLE儿童疾病对照组(包括幼年类风湿关节炎、脊柱关节病、皮肌炎、干燥综合征和血管炎),122例成人SLE和55例其他结缔组织病患者,采用酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)检测血清AnuA。结果 87例儿童SLE中67例AnuA阳性,30例儿童疾病对照组中2例AnuA阳性,AnuA对儿童SLE诊断的敏感性和特异性分别为77.01%和93.33%,ROC曲线下面积为0.852;阳性预测值和阴性预测值分别为97.10%和58.33%。AnuA对成人SLE患者诊断的敏感度和特异度分别为68.85%和90.91%。AnuA对儿童SLE诊断的敏感度高于成人SLE,对儿童SLE诊断的特异度与成人比较无明显差异(P0.05)。在抗dsDNA抗体和抗Sm抗体阴性的儿童SLE患者中AnuA的阳性率分别为56.52%(26/46)和76.71%(56/73)。结论 AnuA对儿童SLE的诊断具有较高的敏感度和特异度,有助于抗dsDNA抗体和抗Sm抗体阴性儿童SLE的诊断。     

Significance of anti-cell membrane-associated DNA (mDNA) antibodies in systemic lupus erythematosus

The aim of this study was to evaluate the value of anti-cell membrane-associated DNA (mDNA) antibodies in the diagnosis of systemic lupus erythematosus (SLE). Antibodies against mDNA were detected with indirect immunofluorescence assay in 207 SLE, 167 other rheumatic diseases, and 82 healthy controls. Association of clinical features and anti-mDNA antibodies was analyzed. The prevalence of anti-mDNA antibodies was 73.3% in SLE, 8.3% in Sjögren’s syndrome, and 4.8% in rheumatoid arthritis. The incidences of anti-mDNA antibodies in SLE lacking antideoxyribonucleoprotein, antihistone antibodies, antinuclesome antibodies, anti-dsDNA, and anti-Sm antibodies were 73.8, 62.7, 65.3, 57.8 and 51.6%, respectively. Skin rash, alopecia, oral ulcer, and joint pain are more common in patients with anti-mDNA antibodies. The anti-mDNA antibody is one of the most valuable markers in SLE. It is also informative in some SLE patients lacking other autoantibodies.     

High frequency of Smith autoantibodies in Omani patients with systemic lupus erythematosus

This study was conducted to investigate the frequency and significance of some antinuclear autoantibodies in Omani patients with systemic lupus erythematosus (SLE). Anti-nuclear antibodies (ANA), anti-double stranded-DNA (anti-dsDNA), and anti-Smith (anti-Sm) autoantibodies were investigated in 60 Omani patients clinically diagnosed with SLE according to the American College of Rheumatology Criteria. The SLE group included 57 females and 3 males with an average age of 26 years. In addition, a group of 60 healthy Omanis (26 females and 34 males; average age 25 years) was used as a control. ANA patterns and autoantibody profile were assayed by indirect immunofluorescence assay using Hep-2 cells and liver/kidney/stomach tissue, respectively. Anti-dsDNA were examined by enzyme-linked immunosorbent assays; anti-Sm antibodies were measured by immunoblotting technique. Out of the 60 SLE patients, 59/60 (98.3%) were seropositive for ANA. Anti-dsDNA and anti-Sm each was detected in 50/60 (83.3%) of the Omani patients. The homogenous pattern of ANA was detected in 30/60 (50%) of patients, whereas the frequency of fine-speckled and coarse-speckled was 16/60 (26.7%) and 6/60 (10%), respectively. High titers (≥1:320) of ANA was detected in 56/60 (93.3%) of SLE patients. High titers of anti-Sm were detected in 22/60 (33.3%) of patients. High titers (>100 IU/ml) of anti-dsDNA were detected in 40/60 (66.7%) of patients. In the control group, ANA were detected in 8/60 (13.3%) but at low titers, whereas anti-dsDNA and anti-Sm were not detected in the healthy control group. This study shows that anti-Sm is as important as the anti-dsDNA for confirming the diagnosis of SLE and that anti-Sm occurs at a much higher frequency (83.3%) than that reported in other populations indicating the importance of this specific autoantibody for the diagnosis and possibly prognosis of Omani SLE patients.     

抗dsDNA抗体两种检测方法的比较

比较抗双链DNA(dsDNA)抗体的两种检测方法。用胶体金快速斑点渗滤技术(DIGFA)及马疫锥虫间接荧光抗体染色法检测27名正常儿童和46例系统性红斑狼疮(SLE)病人和非狼疮病人血清抗dsDNA抗体,27名正常儿童和17例非狼疮病人血清抗dsDNA抗体经两种方法检测均为阴性;29例SLE患者用两种方法检测出抗dsDNA抗体的阳性符合率为94.44％(17/18)。说明两种方法具有一致的特异性和敏感性,DTGFA更简捷、快速、方便。     

Anti-dsDNA antibody avidity determination by a simple reliable ELISA method for SLE diagnosis and monitoring

High avidity anti-dsDNA antibodies are more specific for SLE diagnosis, and more closely associated with renal involvement than intermediate or low-affinity anti-dsDNA antibodies. ELISA methods are largely used to detect anti-dsDNA, but their high sensitivity is inversely related to specificity because they also detect low avidity antibodies. We developed an ELISA assay based on the law of mass action and the competitive binding of dsDNA in solution and coated to microwells with anti-dsDNA antibodies. A simplified Scatchard plot analysis system was used to measure anti-dsDNA antibody avidity which was expressed as apparent affinity constant (Kaa), and quantified in liters per unit (I/U). We prospectively studied 101 consecutive SLE patients, who were followed for 3 years; three serum samples were sequentially collected from each patient during follow-up for determination of IgG anti-dsDNA antibody concentration, and anti-dsDNA avidity. SLE disease activity was estimated using the European Consensus Lupus Activity Measure (ECLAM) index. Sera from 100 healthy subjects and 133 patients with other connective tissue diseases or infectious diseases were also assayed as controls. The mean Kaa in SLE patients was 65.2 +/- 47.3 l/U, with no variations over time. Anti-dsDNA-positive SLE patients had higher Kaa values (79.1 +/- 46.8) than anti-dsDNA negative patients (27.2 +/- 20.1; P < 0.001). No correlation emerged between anti-dsDNA avidity and the ECLAM activity index score. Avidity was significantly higher in patients with renal involvement vs patients without this complication (78.2 +/- 50 vs 59.9 +/- 45.6 l/U; P = 0.0013). This simple ELISA method could be very useful in the diagnostic phase to differentiate high avidity anti-dsDNA autoantibodies that are characteristically found in SLE patients from low avidity antibodies that can also be found in other inflammatory diseases. Moreover, our data confirm the predictive value of high avidity anti-dsDNA antibodies for the development of lupus nephritis.     

Evidence of autoantibodies to cell membrane associated DNA (cultured lymphocytes): a new specific marker for rapid identification of systemic lupus erythematosus

OBJECTIVE—Autoantibodies to cell membrane associated DNA are described in systemic lupus erythematosus (SLE). The specificity of these antibodies differ from antibodies to nuclear DNA.
METHODS—Using indirect immunofluorescence, a specific IgG was detected giving a characteristic pattern of continuous peripheral membrane fluorescence on cultured B-lymphocytes.
RESULTS—This pattern was observed in 53 of 80 serum samples of SLE patients but absent in the serum samples of the control populations: 15 rheumatoid arthritis, 38 ankylosing spondylarthritis, 17 non-inflammatory osteopenic patients, and 224 blood donors. In 34 Sjögren syndrome's patients one only showed a positive test. The cmDNA specificity of these antibodies was confirmed by pattern extinction with DNAse but not RNase or protease pre-treatment of the cells. IgG to cmDNA, separated by absorption/elution from purified cmDNA immobilised on DEAE-nitrocellulose reproduced the immunofluorescence pattern pictures. Extensive serum depletion of anti-double strand or single strand DNA antibodies by absorption to cellulose bound ds- or ss-DNA affected marginally the pericellular fluorescence revealing some minor cross reactivity with nuclear DNA. Moreover, in SLE patients without detectable antibody to ds-DNA, pericellular fluorescence could be visible.
CONCLUSION—This novel rapid immunofluorescence method may serve as an identification test of SLE patients. Given its positive (97.1%) and negative (92.9%) predictive value, sensitivity (66%) and specificity (99.5%), it improves on other diagnostic tests such as the detection of antibodies to Sm.

Keywords: anti-nuclear antibodies; systemic lupus erythematosus, immunofluorescence, cytoplasmic membrane associated DNA     

Anti-Sm: Its predictive value in systemic lupus erythematosus

Summary The clinical manifestations of 131 rheumatic disease patients with anti-Sm antibody were studied. A variety of standard tests was utilized in the study, namely, the FANA test with mouse kidney as substrate for the assay of ANA, the Crithidia test for anti-double stranded DNA (anti-dsDNA) and double immunodiffusion for detecting antibodies to extractable nuclear antigens. The patients were grouped according to the presence of anti-Sm alone, or anti-Sm with some other antibodies. There were 17 with anti-Sm alone; 55 with anti-Sm + anti-RNP; 15 with anti-Sm + anti-dsDNA; and 44 with anti-Sm + anti-RNP. The result of our study showed that although anti-Sm could be found in other diseases, it was exclusively detected in SLE only if anti-dsDNA was also present. Further, the SLE patients with anti-Sm alone had more frequent central nervous system manifestations than other groups of patients. The renal manifestation was observed more frequently in the group of SLE patients with anti-Sm + anti-dsDNA (92.9%). Among other major manifestations, haematologic involvement had a tendency to be less common in the group of patients with anti-Sm alone. The study concludes that the presence of anti-Sm antibody may be of some value to predict the clinical outcome.     

Rapid detection of autoantibodies to dsDNA with the particle gel immunoassay (ID-PaGIA)

OBJECTIVE: To describe a new particle agglutination test for the detection of autoantibodies to double stranded DNA (dsDNA). PATIENTS AND METHODS: Serum samples were collected from 40 unselected healthy blood donors and 200 patients with systemic lupus erythematosus (SLE) or a positive antinuclear antibody screen, or both. The samples were tested in the presence of red high density polystyrene particles coated with purified human dsDNA using the gel technique (Micro Typing System, ID-PaGIA, particle gel immunoassay). The results were compared with those obtained by the two standard anti-dsDNA antibody detection methods, Crithidia luciliae immunofluorescence test (CLIF) and enzyme linked immunosorbent assay (ELISA). RESULTS: The three anti-dsDNA assays exhibited an overall agreement of 87% and significant correlation with each other (p<0.0001). In the SLE group (n=71), 45 patients (63%) were found to be positive by ID-PaGIA compared with only 11/129 (9%) patients in the non-SLE group. Thus the ID-PaGIA had a sensitivity of 63%, and a specificity of 92% for SLE. In comparison, the standard detection methods showed sensitivities of 62% (CLIF) and 70% (ELISA) and specificities of 99% (CLIF) and 84% (ELISA) for SLE. Anti-dsDNA reactivity in the agglutination assay correlated closely with the quantities of antibody obtained by CLIF (r=0.81, p<0.0001) and ELISA (r=0.73, p<0.0001). CONCLUSIONS: The new particle gel agglutination test is a sensitive and specific immunoassay. It is a simple test procedure that might be well suited as a rapid screening method.