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CHARACTERIZATION OF AUTOANTIBODIES IN IMMUNE CYTOPENIAS

免疫细胞减少症中自身抗体的表征

基本信息

批准号：
3363073
负责人：
DENISE R SHAW
金额：
$ 10.32万
依托单位：
UNIVERSITY OF ALABAMA AT BIRMINGHAM
依托单位国家：
美国
项目类别：
财政年份：
1990
资助国家：
美国
起止时间：
1990-07-01 至 1994-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/3363073
关键词：
B lymphocyte affinity chromatography antiidiotype antibody autoantibody autoimmune disorder autoimmune hemolytic anemia enzyme linked immunosorbent assay hemolytic anemia human tissue hybridomas immunogenetics immunoglobulin genes immunoregulation in situ hybridization laboratory mouse monoclonal antibody platelets thrombocytopenic purpura

项目摘要

The etiologies of chronic autoimmune cytopenias like idiopathic thrombocytopenic purpura (AITP) and warm-reactive hemolytic anemia (AIHA) are unknown. This proposal will test the hypothesis that autoantibodies produced in each of these diseases originate from limited repertoires of immunoglobulin variable region germline genes, and that such preferential genetic utilization will be reflected by the presence of frequently shared idiotypes among platelet or red cell reactive autoantibodies from populations of AITP or AIHA patients. The proposed experimental approach is to generate multiple anti-idiotypic reagents, using as immunogens AITP and AIHA autoantibodies generated by two different techniques. The first is to develop human monoclonal autoantibody-secreting cell lines by a novel system: Mice homoxygous for the scid (severe combined immunodefiency) mutation will be stably engrafted with AITP or AIHA patient lymphocytes and immunized with human platelets or red cells, which may activate engrafted human cells but not the genetically unresponsive scid mouse lymphocytes; engrafted and immunized scid mice will be used to generate human monoclonal autoantibody producing hybridoma cells lines. The second approach is direct purification of autoantibodies from AITP and AIHA patients, using antigen-affinity techniques to yield highly enriched polyclonal preparations of platelet or red cell reactive autoantibodies. Both monoclonal and polyclonal autoantibodies will be used to produce BALB/c mouse monoclonal idiotype-specific antibodies. These AITP and AIHA anti-idiotypes will be used in conjunction with extant anti-idiotypes to other human autoantibodies (RF and anti-DNA) to establish the pattern of idiotype expression among multiple AITP or AIHA patients, their family members, and random unrelated unaffected subjects. Additionally, antibody idiotype-positive proteins prepared from various donors will be analyzed for immunoglobulin isotype and binding to relevant autoantigens in vitro, providing data regarding the association of idiotype expression with frank autoimmune disease. Finally, both murine anti-idiotypic reagents and human autoreactive B cell lines generated by the proposed activities will be tools for characterizing specific immunoglobulin genes that encode autoantibodies in AITP and AIHA. These studies will provide important new information regarding genetic parameters in human autoimmune diseases.

慢性自身免疫性血细胞减少症（如特发性）的病因血小板减少性紫癜（AITP）和温反应性溶血性贫血（AIHA）是未知的。这项提议将检验自身抗体这些疾病中的每一种都产生了有限的免疫球蛋白可变区生殖系基因，并且这种优选免疫球蛋白可变区生殖系基因遗传利用将通过经常共享的存在来反映。血小板或红细胞反应性自身抗体中的独特型， AITP或AIHA患者。提出的实验方法以AITP为免疫原，和通过两种不同技术产生的AIHA自身抗体。第一是通过一种新的方法开发人单克隆自身抗体分泌细胞系，系统：scid（严重联合免疫缺陷）同型小鼠突变将稳定地植入AITP或AIHA患者淋巴细胞，用人血小板或红细胞免疫，其可以激活移植的人类细胞，但不是遗传上无反应的scid小鼠淋巴细胞; 移植和免疫的SCID小鼠将用于产生人单克隆抗体产生自身抗体的杂交瘤细胞系。第二种方法是直接纯化来自AITP和AIHA患者的自身抗体，抗原亲和技术，以产生高度富集的多克隆制备血小板或红细胞反应性自身抗体。单克隆和多克隆自身抗体将用于产生 BALB/c小鼠单克隆独特型特异性抗体。这些AITP和AIHA 抗独特型抗体将与现存的抗独特型抗体联合使用，其他人类自身抗体（RF和抗DNA），以建立多个AITP或AIHA患者及其家族中独特型表达成员和随机无关的未受影响的受试者。此外，抗体将分析从不同供体制备的独特型阳性蛋白对于免疫球蛋白同种型和体外与相关自身抗原的结合，提供关于独特型表达与弗兰克相关性的数据，自身免疫性疾病最后，鼠抗独特型试剂和人抗独特型试剂都可以用于抗独特型抗体。通过拟定活动产生的自身反应性B细胞系将用于表征特定免疫球蛋白基因的工具， AITP和AIHA中的自身抗体。这些研究将提供重要的新关于人类自身免疫性疾病的遗传参数的信息。