Detection of anti double-stranded DNA antibodies by RECOMBIGEN anti DNA kit

Anti double-stranded DNA antibodies were detected in patients with various connective tissue diseases by using RECOMBIGEN Anti DNA kit (Japan DPC Co), newly developed utilizing highly purified double-stranded DNA obtained from E. coli plasmid by recombinant technique. High levels of anti double-stranded DNA antibodies were mostly found and distributed in patients with systemic lupus erythematosus and the allied disorders. Because of antibody excess, the final antibody titers could not be determined in most sera which had antibodies titers of more than 100 units per ml. In such sera the expression of antibody titer by DNA binding activities was found to be clinically more useful. RECOMBIGEN Anti DNA kit, thus, is a highly sensitive and reproducible method to detect anti double-stranded DNA antibodies.     

双价抗蛇毒鸡卵黄抗体制备与生物活性研究

目的 通过双价抗蛇毒鸡卵黄抗体(bivalent anti-snake venom immunoglobulin yolk,双价IgY)的制备及其相关特性研究,为多价抗蛇毒IgY的制备和应用奠定基础.方法 两种单一抗原(舟山眼镜蛇、圆斑蝰泰国亚种)按顺序依次交替注入单只鸡体内,水稀释法制备双价IgY;测定双价IgY效价(间接ELISA法)、交叉免疫特性(双向免疫扩散试验)及对溶膜活性(溶膜试验)、半数致死量(LD50)的中和作用.结果 初次免疫后第28-42天,以水稀释法提取的双价IgY对眼镜蛇毒及蝰蛇毒的效价分别为1:12 800和1:6400.双价IgY与眼镜蛇亚科、蝰业科6种蛇毒有明显的交叉免疫反应,而与蝮亚科4种蛇毒的交叉免疫反应不明显.双价IgY能显著降低眼镜蛇、蝰蛇毒对鸡卵黄膜的溶膜作用,也能显著延长眼镜蛇和蝰蛇伤小鼠的存活时间(P<0.05),同等剂量的双价lgY提高眼镜蛇伤存活率优于蝰蛇伤.结论 双价IgY能够显著中和眼镜蛇、蝰蛇毒的溶膜和致死活性,能有效保护机体免受眼镜蛇毒和蝰蛇毒的攻击.     

抗HBsAg和抗红细胞双特异Diabody的构建及表达

目的构建及表达抗乙肝病毒表面抗原（ＨＢｓＡｇ）和抗红细胞（ＲＢＣ）双特异Ｄｉａｂｏｄｙ，用于血清中ＨＢｓＡｇ的快速检测。方法将抗ＨＢｓＡｇ的人抗体轻重链可变区基因与抗红细胞的鼠抗体重轻链可变区基因交叉配对构建成两个杂合Ｄｉａｂｏｄｙ基因，并将两个配对基因组建在一个表达载体中，成为双特异Ｄｉａｂｏｄｙ表达载体，并在大肠杆菌中分泌表达。结果经ＥＬＩＳＡ检测和红细胞凝集实验测定，证明所表达的双特异性Ｄｉａｂｏｄｙ具有抗ＨＢｓＡｇ和抗ＲＢＣ双重功能，并可使含有ＨＢｓＡｇ的ＲＢＣ悬液产生血球凝集。结论所表达的双特异性Ｄｉａｂｏｄｙ具有双重抗体活性，可用于血清中ＨＢｓＡｇ的快速检测     

IgG型抗CD55×抗β-Gal基因工程双特异性抗体的构建

目的　探讨用生物学方法治疗病毒感染性疾病及肿瘤的新途径。方法　以有重要补体活化调节功能的膜补体调节蛋白CD5 5为靶点 ,以 β GaL为模拟病毒或肿瘤抗原 ,制备“IgG”型抗CD5 5×抗 β Gal基因工程双特异性抗体 ,并对该重组抗体真核表达后的结合活性进行初步的验证。结果　克隆的CD5 5抗体可变区片段为新的小鼠抗体可变区片段 ,经HEK 2 93细胞表达后的重组抗体显示了良好的CD5 5及Fc结合活性。结论　本研究为病毒性疾病或肿瘤的免疫学治疗提供了新的途径及实验依据     

抗—HBs基因水平的研究

本研究应用ＲＴ－ＰＣＲ、ＤＮＡ－ＰＣＲ及Ｓｏｕｔｈｅｒｎｂｌｏｔ等分子生物学技术检测不同人群中针对一株抗－ＨＢｓ优势克隆的基因水平。发现慢性乙肝（ＨＢｓＡｇ阳性）患者中检出率为２０．６９％（６／２９），明显低于乙肝疫苗注射后人群的６２．９６％（１７／２７），Ｐ＜０．０１；而与未接种过乙肝疫苗组的检出率相比则无明显差异（Ｐ＞０．０５），后者阳性率为２１．４２％。这一结果提示：慢性乙肝患者与乙肝疫苗注射后相比，在抗－ＨＢｓ基因水平上存在差异，可能有抗－ＨＢｓ克隆的缺失或转录、ｍＲＮＡ翻译合成抗－ＨＢｓ的异常。本工作中对人群中抗－ＨＢｓ基因研究的尝试，为乙肝免疫耐受的研究提供了新的线索，同时也为乙肝的基因治疗提供了一定依据。     

对两种第三代丙肝试剂检测的不同功能区抗体组份研究

目的　比较第二代和第三代丙肝试剂对丙肝不同功能区抗体的检出能力。方法　用两种第二代抗 Ｈ Ｃ Ｖ Ｅ Ｉ Ａ 试剂（ Ｋ２ 中国制造, Ａ２ 美国制造） 及两种第三代主流试剂（ Ａ３ 德国制造, Ｏ３ 美国制造） 对１２１ 份来源于全国各地的供血员血样进行了检测。结果　该１２１ 份样品经 Ｒ Ｉ Ｂ Ａ Ｈ Ｃ Ｖ３ ．０ 试剂及 Ｐ Ｃ Ｒ 方法确证, 其中４３ 份为阳性样品,７８ 份为阴性样品。两种第二代试剂 Ｋ２ , Ａ２阳性检出符合率为３６／４３ 和３７／４３ , 两种第三代试剂 Ａ３ , Ｏ３ 阳性检出符合率分别为４０／４３ 和３９／４３ 。在４３ 份阳性样品中,２０ 份既含有抗 Ｈ Ｃ Ｖ 核心抗体, 也含有抗 Ｈ Ｃ Ｖ Ｎ Ｓ３ 抗体,用 Ｋ２ , Ａ２ , Ａ３ , Ｏ３ 四种试剂分别检出１８ 、１８ 、１９ 和２０ 份。１４ 份含有抗 Ｈ Ｃ Ｖ Ｎ Ｓ３ 抗体而不含有抗 Ｈ Ｃ Ｖ 核心抗体的样品中,用两种第二代试剂（ Ｋ２ , Ａ２） 分别检出９ 份和１０ 份, 而用两种第三代试剂（ Ａ３ , Ｏ３） 则全部检出。与此相反,９ 份含有抗 Ｈ Ｃ Ｖ 核心抗体而不含抗 Ｈ Ｃ Ｖ Ｎ Ｓ３ 抗体的样品, 用两种第二代试剂（ Ｋ２ , Ａ２）均可检出, 而用两种第三代试剂（ Ａ３ , Ｏ３） 却     

Anti‐idiotype and anti‐anti‐idiotype antibodies for aflatoxin from laying hens

Anti‐idiotype (anti‐id) antibodies (IgY2)for aflatoxin (AF) were obtained from the egg yolks of laying hens immunized with affinity‐purified rabbit polyclonal anti‐aflatoxin B₁ (AFB₁) carboxymethyloxime‐bovine serum albumin (BSA) antibodies (pAb1). The IgY2 were affinity purified and then subjected to various analyses. Inhibition of the binding of pAbl to the solid‐phase AFB₁‐BSA by IgY2 and the binding of pAb1 to the solid‐phase IgY2 by free AFB₁ were demonstrated in a biotin‐avidin amplified enzyme‐linked immunosorbent assay (ELISA) system. The concentration of IgY2 causing 50% inhibition (ID₅₀) of the binding of pAb1 to AFB₁‐BSA was found to be 2.45 μg/assay. The ID₅₀ concentration of the binding of pAb1 to IgY2 by free AFB₁ was found to be 0.30 μg/assay. Inhibition of the binding of AFB₁‐horseradish peroxidase (HRP) to the solid‐phase pAbl by IgY2 (ID50 = 9.65 μg/assay) was also demonstrated in the direct ELISA. Egg yolk anti‐anti‐id antibodies (IgY3) were obtained by immunizing laying hens with rabbit pAb2 against anti‐AFB₃‐hemisuccinate‐BSA monoclonal antibody. IgY3 was subjected to affinity chro‐matography purification with Sepharose gel armed with AFB₂‐carboxymethytoxime, and then subjected to various analyses. ELISA analysis revealed that IgY3 has characteristics similar to other anti‐AFB antibodies induced in various experimental animals. In the direct ELISA, the ID₅₀ of the binding of AFB₁‐HRP to solid‐phase lgY3 by AFB₁ was found to be 0.12 ng ml^‐1. In the indirect ELISA, the ID₅₀ of the binding of IgY3 to solid‐phase AFB₁‐BSA by AFB₁ was found to be 2.2 ng ml^‐1. The IgY3‐based ELISA analysis showed higher sensitivity than that of the egg yolk antibodies directly against AFB‐protein conjugates (IgY1). A good correlation was found for the data obtained from IgY3‐based and pAb1‐based ELISAs in the analysis of AFB in the fungal culture filtrates.     

Hemolytic disease of the newborn due to anti S antibodies

We report a case of hemolytic disease in a newborn due to anti S antibodies. Baby R was born at term to an O+ mother whose antibody screen was positive for phenotype big S. Cord blood eluate revealed anti-S RBC; antigen: RBC typing for S- was positive. Physical examination of baby was unremarkable. The infant's HCT was 44.2 at 6 hours of age. At 48 hours, the HCT decreased to 33.5, bilirubin peaked to 5.4, retic had peaked to 6.8. By seven days, all these values reverted to the normal, and baby has remained asymptomatic.     

抗红细胞双价小分子抗体的构建及表达

单链抗体（ＳｃＦｖ）是一个重链可变区（ＶＨ），一个轻链可变区（ＶＬ）由连接肽（Ｌｉｎｋｅｒ）连在一起构成的单价小分子抗体。为使其能组装成双价，对一个抗人红细胞的单链抗体进行了改造，将其连接肽由１７个氨基酸〔ＳＲ（ＧＧＧＧＳ）３〕缩短为６个氨基酸（ＳＲＧＧＧＳ），强迫不同分子间的ＶＨ和ＶＬ组合成Ｆｖ，从而形成双价小分子抗体（Ｄｉａｂｏｄｙ）。在大肠杆菌中分泌型表达后，显示具有血球凝集活性，证明其为双价。进一步用凝胶过滤分析，显示改建后抗体分子的分子量约为原单链抗体的两倍。     

Evaluation of IgM anti human cytomegalovirus antibody

An anti-human cytomegalovirus (HCMV) antibody in 330 sera from patients and normal subjects was examined by ELISA. The test results were compared with the results of CF test and there was 99.1% agreement. Specific anti-HCMV IgM antibody was searched for and only 4 sera were positive. This makes it unlikely that screening for IgM anti-HCMV will effectively prevent posttransfusion HCMV infections.     

人抗动物抗体及其对免疫分析的干扰

人抗动物抗体可引起临床不良反应，干扰动物源性抗体制剂治疗或成像，干扰免疫分析而引起假性结果。临床上可通过应用免疫抑制剂治疗和人源性抗体、聚乙二醇处理的抗体或体Ｆａｂ段制剂等方法防止人抗动物抗体应答形成；样品预处理或分析技术的改进可消除人抗动物抗体对免疫分析的干扰。目前已设计出了多种分析技术检测人抗动物抗体，并形成了检测试剂盒，甚至床边检验试剂。     

重组抗HBx单链噬菌体的表达

目的：进一步探索重组基因工程抗体在原核系统中有效的活性表达途径。方法：在已构建的抗ＨＢｘ单链抗体（ＳｃＦｖ）的基础上,利用粘粒载体构建了抗ＨＢｘ单链噬菌体抗体（ｐｈａｇｅ ａｎｔｉｂｏｄｙ）,并转化大肠杆菌。结果：该噬菌体抗体经辅助噬菌体Ｍ３Ｋ０７诱导和噬菌体次要衣壳蛋白（ＰⅢ）融合表达,并组装成噬菌体衣壳蛋白可溶性表达于培养液上清,经ＥＬＩＳＡ和Ｗｅｓｔａｒｎ－ｂｌｏｔ的结果证实可溶性表达产物具     

医学知识管理平台的开发及应用

医疗服务行业是知识密集型行业,医疗技术和管理水平构成了一家医疗机构的核心竞争力.医疗服务行业的竞争实质就是医疗技术和管理水平的竞争.通过计算机技术的应用,实现医学知识管理的目标,从而提高医院的核心竞争力,有大量相关的管理和技术问题需要讨论.     

Detection of anti rotavirus coproantibodies by immunoblotting technique

IgA and IgG coproantibodies to individual simian rotavirus (SA 11) structural polypeptides were detected in healthy infants in nursery homes. The number of immunoblottable peptides differed from individual to individual. Coproantibodies were also detected at the convalescent stage of rotavirus infection in two patients but not during the acute stage. This method is useful for confirming the diagnosis of rotavirus infection serologically without the need for paired sera.     

Anti‐idiotype and anti‐anti‐idiotype antibodies generated from polyclonal antibodies against aflatoxin b1

Polyclonal anti‐idiotype antibodies (anti‐id, pAb2) for aflatoxin were generated from mice ascites after immunization with affinity‐purified rabbit polyclonal antibody (pAbl) against aflatoxin B₁ (pAFB₁). After affinity chromatography purification, pAb2 were subjected to various analyses, and were used to generate anti‐anti‐id antibodies (pAb3) in the ascites of Balb/c mice. ELISA analyses revealed that the anti‐id antibodies bound specifically to the original pAb1, but not to other types of monoclonal antibodies or normal mouse IgG. The inhibition of binding of AFB₁ to pAb1 by pAb2 was demonstrated in both regular and biotin‐avidin amplified enzyme‐linked immunosorbent assay (ELISA) systems in which AFB₁‐bovine serum albumin (BSA) was coated on to the microtiter plate. The concentrations causing 50% inhibition (ID₅₀) of the binding of pAbl to AFB₁‐BSA by pAb2 were found to be 6.5 and 1.7 μg/assay respectively. Inhibition of the binding of pAb1 to the solid‐phase pAb2 by free AFB₁ (ID₅₀ = 0.63 μ/assay) was also demonstrated in the ELISA. pAb3 were found to have similar characteristics to the original pAb1. In the direct ELISA, the ID₅₀ of binding of AFB₁‐horseradish peroxidase to the solid‐phase pAb3 by AFB₁ was found to be 0.20 ng ml^‐1. In the indirect ELISA, the ID₅₀ of the binding of pAb3 to the solid‐phase AFB₁‐BSA by AFB₁ was found to be 1.84 ng ml^‐1 Analysis of a naturally contaminated corn sample for AFB₁ by the pAb3‐based ELISA gave a result similar to that obtained from the regular pAb1‐based ELISA (19.7 and 21.8 μg kg^‐1 respectively).     

Naturally occurring anti M complicating ABO grouping

Anti M is considered a naturally occurring antibody that is usually active at temperatures below 37°C and is thus of no clinical significance. This antibody, if present in an individual, can lead to a discrepancy between forward and reverse ABO grouping and thus creates diagnostic difficulties for blood bank staff. We report a case of a 58-year-old lady who had an unexpected reaction in reverse grouping due to anti M that posed a problem for us in the interpretation of results of her blood group. We also reviewed the literature to find out the significance of such discrepancy in blood grouping.     

Mycophenolate mofetil and roscovitine decrease cyclin expression and increase p27(kip1) expression in anti Thy1 mesangial proliferative nephritis

The response of mesangial cells to a phlogistic challenge includes cell proliferation and mesangial matrix expansion. Cell proliferation is a highly regulated process which includes enhancing factors such as cyclins, cyclin dependent kinases, and inhibitory proteins, such as p27(kip1). The aim of the study was to evaluate the effects of Mycophenolate mofetil (MMF), and roscovitine (R), on the cell cycle regulatory system when administered in the florid phase of the experimental model of mesangial proliferative nephritis induced by the anti Thy-1 antigen monoclonal antibody. Three days after nephritis induction, different groups were given MMF and R. Rats treated with MMF or R showed a slight decrease in mesangial proliferation and matrix expansion. Samples of cortical tissue were tested by 'real time' RT-PCR in order to study gene expression of cyclins B, D1, D2, D3, E, and the cyclin inhibitor p27(kip1). Localization of mRNA was evaluated by in situ hybridization. Real time RT-PCR analysis showed a significant decrease in cyclins B, D1, D2, and D3 in rats treated with either MMF or R as compared to controls. Both MMF and R treatment induced a significant increase in p27(kip1) mRNA expression. In situ hybridization showed a mesangial-endothelial expression pattern in glomeruli. The number of labelled cells per glomerulus, the number of positive glomeruli in each examined slide as well as cyclin D2 and D3 signal intensity was significantly lower in rats treated with MMF or R as compared to controls, whereas MMF or R treatment up-regulated p27(kip1) mRNA expression. Immunohistochemical evaluation of p27(kip1) aimed to examine the influence of MMF or R on protein expression confirmed up-regulation.