ABC法与ELISA法诊断人体包虫病的比较研究

本文应用ABC-ELISA法对101例经手术证实的包虫病人,40例非包虫病患者及61例正常人进行了血清抗细粒棘球蚴(E.g.)特异性抗体检测,并同时行常规ELISA法进行比较。101例包虫病人中ABC法和ELISA法与术后诊断的阳性符合率分别为94.06％和81.19％,对照组假阳性率分别为1.98％和0.99％。两法同为阳性的包虫病人中ABC法检出的抗体滴度为ELISA法的6.70倍。13例ELISA法阴性的包虫病人ABC法阳性,提示ABC法对于那些抗体水平较低的包虫病人可能更具有诊断价值。本实验结果表明:ABC法诊断人体包虫病较ELISA法敏感性高,且具有特异性好、简便、快速的优点。     

抗体捕获ELISA检测包虫病人特异性IgM抗体及其应用的研究

用标记抗原、标记抗体、标记抗抗体及ABC四种形式的抗体捕获ELISA法检测包虫病人血清中特异性IgM抗体,结果以ABC-捕获ELISA法阳性率最高为52.5％,标记抗抗体法次之为39.6％,标记抗体法为22.5％,标记抗原法阳性率最低为20.7％。四种方法检测健康人血清均为阴性。手术前的包虫病人血清中特异性IgM抗体的阳性率明显高于手术后者,分别为52,2％和20％。不同寄生部位包虫病人特异性IgM抗体的阳性率没有明显差别。在入院就诊的病人中阳性率为35.4％,而在普查中发现的无症状病人中阳性率达70.5％,说明特异性IgM的检测在包虫病中也具有早期诊断的意义。     

人体包虫病主要特异性免疫球蛋白的水平及其临床意义

本文报道了应用酶联免疫吸附试验(ELISA)检测包虫病人血清中抗包虫特异性免疫球蛋白(IgG和IgM)的水平并对其临床意义进行了初步探讨。48例正常人血清特异性IgG和IgM的假阳性检出串均为0％;24例囊尾蚴病人血清特异性抗包虫IgG和IgM的假阳性检出效分别为:33.4％;4.16％;42例包虫病患者血清特异性IgG和IgM的阳性检出率分别为:80.95％,66.67％;两者之差经统计学处理具有显著意义(p<0.05)。肝包虫特异性IgG的阳性检出率较肺包虫高,肺包虫特异性IgM的阳性检出率较肝包虫高。这提示患者对包虫的免疫应答和其寄生的位置有关;诱导产生的特异性免疫球蛋白主要为IgG类,特异性IgM的水平与包虫囊壁的完整与否以及检测方法的灵敏性、特异性有关。     

包虫抗原的免疫亲和层析纯化与鉴定

<正> 包虫(细粒棘球蚴Echinococcusgranulosus,Eg)囊液(CF)为包虫病最常用的免疫诊断抗原。在ELISA试验中,以粗囊液作为抗原,用量少则敏感性差;用量大却非特异反应重,故纯化包虫抗原显得尤为重要。我们以包虫病人血清Ig交联的免疫亲和层析法纯化包虫抗原,经ELISA及免疫印渍法与半饱和硫酸铵盐析纯化的EgCF抗原及粗EgCF抗原作对比,结果如下。     

抗包虫囊液抗原单克隆抗体细胞系的建立及临床应用研究

本文用SP2/0小鼠骨髓瘤细胞与人肺细粒棘球绦虫(E.granulosus)囊液免疫BALB/C小鼠,取其脾脏进行细胞融合、获得了稳定分泌IgG Ⅰ型抗包囊虫的杂交瘤细胞系。染色体数为95—105条,腹水ELISA效价为320000的倒数,用该抗体建立了双抗ELISA夹心法,对包虫疫区—新疆和布克赛尔蒙古自治县5000人采用耳垂血沪纸片浸液检查,阳性率为10.62％,与B型超声波检查附合率为97.63％、与间接ELISA相比有较高的特异性、大大缩短了检查时间,适合基层开展包虫病的普查。     

包虫病特异性抗体检测试剂盒诊断效能评价及结果判定优化

为评价3种包虫病特异性IgG抗体诊断试剂盒的临床诊断效能,并针对其特异度差的问题进行优化,为包虫病免疫学诊断提供检测结果优化判定的策略,本研究共收集87例包虫病确诊病人血清,78例经排查为非包虫病就诊病人血清.每份血清同时采用3种试剂盒进行检测,对结果进行统计分析,评价各方法的诊断效能.通过受试者工作特征(ROC)曲线寻找各方法最佳诊断界值.采用胶体金免疫渗滤斑点法(DIGFA)对酶联免疫吸附法(ELISA)或胶体金免疫层析法(GICA)所得检测结果低于最佳诊断界值的阳性样本进行重新测定,以DIGFA法最佳诊断界值为结果判定标准.比较结果优化前后的诊断效能.结果 显示,3种诊断方法的灵敏度在90.8％ ～94.3％之间,特异度在75.7％～83.3％之间.ROC曲线所得最佳诊断界值均高于试剂盒的cutoff值.检测结果经优化后,灵敏度保持在90.3％以上,特异度提升至88.5％以上.结果 表明,3种诊断方法的特异度均不理想,本次研究通过对检测结果的优化判定,在不影响灵敏度的前提下可减少该部分血清假阳性结果的误判,提高方法的特异度,为包虫病临床诊断提供更为可靠的信息.     

包虫病患者血清中循环免疫复合物测定及其临床意义的探讨

<正> 包虫病患者血清中循环免疫复合物(Circulating Immune Complexes,CIC)升高已为一些学者的研究所证实。但其临床意义尚不能肯定。本文测定了62例确诊为包虫病的患者血清中CIC浓度,同时检测了其血清中抗细粒棘球蚴(Echinococcus granulosus,E.g)抗体滴度,旨在了解包虫病人血清中CIC水平,探讨CIC与包虫囊肿临床表现及血清中抗E.g.特异性抗体水平之间的关系。     

ELISA和ABC免疫组化染色法在肿瘤McAbs筛选中的比较研究

本文比较分析了ELISA和ABC两种方法在肿瘤McAbS筛选应用中的一些利弊关系。结果表明,ELISA测出为阳性并与结肠癌细胞系反应的99.5％(198/199)孔杂交瘤细胞所分泌的抗体,经ABC法染色后是不与其癌组织反应的,其中只有0.5％(1/199)孔细胞的抗体才是这两种方法同时均能测得为阳性和是有意义的。而ABC法不仅可检出ELISA阳性的抗体,而且亦能检出ELISA漏筛为性阴、最终是有意义的理想抗体。     

点免疫金渗滤法筛选胶体金标记抗人IgG 的研究

目的:应用点免疫金渗滤法筛选适宜胶体金标记的抗人IgG(以下简称二抗),并运用多个评价指标进行比较和分析。方法:选择三个不同公司的二抗用胶体金标记,根据棋盘滴定法确定最佳标记条件;运用点免疫金渗滤法对三种二抗的胶体金标记后检测效能进行比较,采用包虫病人和健康对照血清,用包虫病特异性抗原检测包虫病患者血清中的特异性抗体,评价最优的二抗,并与酶联免疫吸附测定法进行比较。结果:A、B、C 三种二抗标记的最佳标记条件为:pH 均为8.5,加入量分别为38.4、24、19.2 g/ ml,综合评价二抗B 检测效能最优。将其与酶联免疫吸附测定法检测效能进行比较,两种方法检测结果的Kappa=0.895(P<0.05),吻合度较强。结论:应用点免疫金渗滤法,以及本文提出的评价指标,能够筛选出最佳二抗,对检测试剂盒中二抗的选择具有重要的参考价值。     

用ABC-ELISA提高检测脑脊液结核抗体的敏感性

本文用ABC—ELISA检测40例结核性脑膜炎及66例对照患者CSF中特异性IgG抗体,并在同一滴定板上做常规ELISA对比。结果表明ABC—ELISA检测CSF抗体滴度高于常规ELISA的3倍;抗体阳性的检出时间ABC法也早于常规法1天;其阳性检出率也明显高于常规法(p<0.05),而其非特异性未见增加,提示本法可能为结核性脑膜炎的免疫学诊断提供一种比常规ELISA更为敏感的新方法,有助于低抗体水平患者的检出。     

Enzyme-linked immunosorbent assay for immunoglobulin M specific antibody for the diagnosis of melioidosis.

Indirect hemagglutination (IHA) is commonly used for serodiagnosis of melioidosis. However, in endemic areas, high background titers in normal populations and occasional low titers in patients with septicemic melioidosis prompted a search for a more sensitive and more specific method of serodiagnosis. An indirect fluorescent-antibody test for immunoglobulin M (IgM) specific antibody to Pseudomonas pseudomallei was more sensitive and more specific, but fluorescence microscopes are rarely available in the endemic areas. An enzyme-linked immunosorbent assay (ELISA) for IgM antibody is an attractive alternative. An indirect ELISA for IgM antibody (IgM ELISA) and an IgM antibody capture ELISA for melioidosis were developed. Both tests, together with IHA, were evaluated for 153 serum specimens from blood donors and 16 serum specimens from 16 melioidosis patients. It was found that IHA, the IgM ELISA, and the IgM antibody capture ELISA had sensitivities of 88, 88, and 75%, respectively, with specificities of 97.4, 92.2, and 91.5%, respectively. When IHA was combined with IgM ELISA, a sensitivity of 100% and a specificity of 95.4% were obtained. The IgM ELISA and IHA should be used in combination for serodiagnosis of melioidosis.     

Evaluation of an enzyme-linked immunosorbent assay that uses ferrous metal beads for determination of antihistoplasmal immunoglobulins G and M

A rapid enzyme-linked immunosorbent assay (ELISA) for the detection of human class-specific antibodies to Histoplasma capsulatum (histoplasmal immunoglobulin M [HIgM] and histoplasmal IgG [HIgG]) was developed by using antigen adsorbed onto polycarbonate-coated ferrous beads. In the ELISA method all the reagents used were commercially available. In 135 specimens from patients with confirmed histoplasmosis, sensitivities were 76% for complement fixation (CF), 53% for immunodiffusion (ID), and 64% for the ELISA for HIgM and HIgG combined. The ELISA detected histoplasmal antibody in 36% of the specimens with negative antibody titers by CF and 46% of the specimens with negative antibody titers by ID. The ELISA detected histoplasmal antibody in 27% of specimens that were negative by both CF and ID. When limited to specimens collected within 4 months of the onset of histoplasmosis symptoms, sensitivities were 82% for CF, 63% for ID, and 86% for ELISA for HIgG and HIgM combined. Within this group, ELISA detected histoplasmal antibody in 90% of the specimens that were negative by CF, 76% that were negative by ID, and 100% that were negative by both CF and ID. The specificity of the ELISA could not be fully addressed since sera from patients with other fungal infections were not available.     

Prevalence of Aspergillosis in chronic lung diseases

Eighty eight patients of chronic lung diseases (CLD) attending TB and Chest department of J.N. Medical college Hospital were studied to find out the prevalence of Aspergillus in Broncho-alveolar Lavage (BAL) and anti- aspergillus antibodies in their sera. Direct microscopy and fungal culture of BAL was done. Antibodies were studied by immunodiffusion (ID) and Enzyme linked immunosorbent assay (ELISA). Dot blot assay for anti-aspergillus antibodies was also performed in sera of patients which were either positive by ID or by ELISA. Aspergillus was isolated in culture from 13(14.7%) cases of CLD, while, 30.6% cases showed anti-aspergillus antibodies by serological methods. Aspergillus fumigatus was the predominant species isolated. 17(19.3%) cases of CLD showed antibody against Aspergillus by ID, 22(25%) by ELISA, while 19 of 27 seropositive cases also showed positive results by Dot Blot assay. In cases of bronchogenic carcinoma and pulmonary tuberculosis, anti-aspergillus antibodies were detected equally by ID and ELISA in 21.42% and 21.05% cases respectively. In bronchial asthma, the antibodies could be detected in 60% cases by ELISA, while, in only 10% cases by ID. ELISA was found more sensitive than ID for detection of anti-aspergillus antibodies. The sensitivity of Dot Blot lies some what between ID and ELISA. It is concluded that prevalence of Aspergillosis is quite high in chronic lung diseases, culture and serological test should be performed in conjunction and more than one type of serological tests should be performed to establish the diagnosis.     

Circulating immune complexes in sera of patients infected with Echinococcus granulosus

Circulating immune complexes (CIC) were investigated by the C1q binding assay in sera of 23 patients infected with Echinococcus granulosus. For the 23 sera studied, nine were found to be positive in the test. When the samples were grouped according to the cyst localization, the highest rate of CIC positively was found in the group of sera from patients with pulmonary cyst; in this group, double diffusion (DD) and indirect haemagglutination (IHA) tests gave a low rate of positivity for antibodies directed against parasitic antigens. Rheumatoid factor, anti-nuclear and anti-mitochondrial autoantibodies were not detectable in patients' sera by indirect immunofluorescence technique (IIF). Anti-smooth muscle autoantibodies, detectable by IIF, were present in 56% of the sera and this positivity was higher in the hepatic (83%) than in the pulmonary form (40%).     

应用生物素-亲和素系统检测胸水特异性IgG抗体的研究

应用亲和素—生物素化过氧化物酶复合物酶联免疫吸附试验(ABC-ELISA)检测159份结核性胸水和82份对照胸水中的抗PPD—IgG,并在同一滴定板上做常规ELISA对比。结果表明在82.4％的胸水中,ABC—ELISA较常规ELISA敏感,前者的抗体几何平均滴度为后者的2.4倍。本法敏感性为90.6％,特异性为95.1％,提示可作为结核性胸膜炎的辅助诊断方法。动态观察尚可为疗效判定提供依据。     

Serodiagnosis of imported schistosomiasis by a combination of a commercial indirect hemagglutination test with Schistosoma mansoni adult worm antigens and an enzyme-linked immunosorbent assay with S. mansoni egg antigens

A commercial indirect hemagglutination (IHA) test using erythrocytes coated with Schistosoma mansoni adult worm antigens (WA) and an enzyme-linked immunosorbent assay (ELISA) with S. mansoni egg antigens (SEA) were assessed for their use in serodiagnosis of imported schistosomiasis (hereafter these tests are designated WA/IHA and SEA/ELISA, respectively). The sensitivity of the tests was evaluated with sera from 75 patients with proven S. mansoni infection, 25 with proven S. haematobium infection, and 10 with clinical Katayama fever. The specificity was assessed with sera from 283 patients with various parasitic, bacterial, viral, and fungal infections and sera containing autoimmune antibodies. Sensitivities of the WA/IHA with a cutoff titer of 1:160 (WA/IHA(160)) in detecting S. mansoni, S. haematobium, S. mansoni and S. haematobium combined, and clinical Katayama fever were 88.0, 80.0, 86.0, and 70.0%, respectively, with a specificity of 98.9%. The WA/IHA with a cutoff of 1:80 (WA/IHA(80)) showed sensitivities of 94.7, 92.0, 94.0, and 90.0%, respectively, with a specificity of 94.7%. The comparable values of SEA/ELISA were 93.3, 92.0, 93.0, and 50.0%, respectively, with a specificity of 98.2%. Combined use of ELISA and WA/IHA(80) gave sensitivities of 100% for S. mansoni, S. haematobium, and S. mansoni and S. haematobium combined and 90% for Katayama fever. The specificity of this combination in detecting schistosomiasis was 92.9%. Combination of SEA/ELISA with WA/IHA(160) gave sensitivities of 98.7, 96.0, 98.0, and 80% with a specificity of 97.2%. Our findings suggest that WA/IHA and SEA/ELISA are each sensitive and specific serological tests that are easy to use for the diagnosis of imported schistosomiasis. The combined use of these two tests enabled the serological diagnosis of schistosomiasis to be achieved with very high degrees of both sensitivity and specificity.     

Cysticercus immunoblot assay in patients with single, small enhancing lesions and multilesional neurocysticercosis

A single, small (< 20 mm), ring or disc shaped contrast enhancing lesion located at the cortical-subcortical junction with minimal or no surrounding edema on computed tomography is the commonest mode of presentation of neurocysticercosis in the Indian subcontinent. Serum samples of 37 patients with these single, small enhancing lesions (SSEL's) and five patients with typical multilesional parenchymal neurocysticercosis were tested by the electro-immunoblot transfer (EITB) assay and the enzyme linked immunosorbent assay (ELISA). EITB was positive in 18 patients (48.64%) and ELISA was positive in 21 patients (56.76%) with SSEL's. On the other hand EITB was positive in all five patients (100%) and ELISA was positive in four patients (80%) with multilesional neurocysticercosis. The low sensitivity of the EITB in the SSEL's is probably linked to an insufficient immune stimulation provided by a single cysticercus cyst.     

Prevalence of aspergillosis in bronchogenic carcinoma

Bronchoalveolar lavage of 42 patients of bronchogenic carcinoma was studied to find out the prevalence of aspergillosis. Sera of the patients were also analysed for presence of anti-Aspergillus antibodies by Immunodiffusion (ID), Enzyme linked immunosorbent assay (ELISA) and dot blot assay (DBA). Aspergillus was isolated in culture from 6 (14.2%) patients of bronchogenic carcinoma. Aspergillus fumigatus was the predominant species isolated. All the strains of Aspergillus were sensitive to itraconazole, ketoconazole and amphotericin B while resistance (33.3%) was found with fluconazole. Anti-aspergillus antibodies were detected equally by ID, ELISA and DBA in 9 (21.4%) cases. The present study revealed prevalence and seroprevalance of Aspergillus in bronchogenic carcinoma to be 14.2% and 21.4% respectively. Consistent reactivity against 18 kDa Aspergillus fumigatus antigen was noted in serologically positive cases. Antibodies against 18 kDa protein antigen in western blotting may be used as a reference marker for diagnosis of aspergillosis in bronchogenic carcinoma. It is also suggested that the simplest serological technique like ID may be performed along with culture for diagnosing Aspergillosis in patients of bronchogenic carcinoma since ID, ELISA and DBA showed similar sensitivity.     

High-quality, cost-effective strategy for detection of autoantibodies to extractable nuclear antigens

We evaluated methods for the detection of autoantibodies to extractable nuclear antigens (ENAs) to determine the strategy that yielded the most cost effective and clinically meaningful result. We prospectively compared counterimmunoelectrophoresis (CIEP) with and without serum prediffusion (SPD) and found that SPD significantly improved the quality of precipitation lines. This resulted in a decreased requirement for repeat testing and, consequently, was associated with a significant decrease in reagent costs and specimen turnaround time. We also retrospectively compared reactivity by CIEP, CIEP plus SPD, enzyme-linked immunosorbent assay (ELISA), and line immunoassay (LIA) of 52 serum samples that were previously determined to be positive for ENAs, and we correlated the results with clinical diagnoses. There was significant agreement among CIEP, CIEP plus SPD, ELISA, and LIA for the detection of anti-SS-A, anti-SS-B and anti-RNP. In general, CIEP, CIEP plus SPD, and LIA correlated better with the clinical diagnoses than ELISA, even though ELISA detected anti-ENAs more often than the other methods. CIEP plus SPD is therefore the most cost effective method for the identification of clinically meaningful ENAs. Based on our experience, we now screen for ENAs by CIEP, and positive samples are then typed by CIEP plus SPD. Samples that are difficult to interpret are then further assessed by an alternative method.