DEVELOPMENT OF DIAGNOSTIC PROBES FOR AUTOIMMUNE ANTIGENS

许多风湿性疾病的一个特征是 存在循环抗核抗体。 试验基于 这些抗体的测量建立在 临床实验室诊断这些疾病，但往往 如果没有很好地表征或分离 靶抗原。 一种这样的自身免疫性疾病是硬皮病或 进行性系统性硬化症 本提案的目的是 提供灵敏和定量的诊断/预后测定 通过开发基于直接结合的检测方法来治疗这种疾病 重组多肽抗原的血清抗体。 的 最初的重点将是用该方法鉴定的五种多肽。 两种主要的硬皮病相关抗原，SCL-70和 着丝粒。 抗原将以足够的量分离 用于微测序。 基于这些的寡核苷酸探针 序列将被用来克隆这些基因编码 来自人cDNA文库的多肽。 表达 克隆的基因将允许合成克量的 抗原足以进行测定。 的 重组多肽也可用作免疫原 确保抗原抗体恒定和充足的供应 分离和最终测定标准化。 类似的努力 将被导向其他与硬皮病相关的 抗原，包括原纤蛋白、PM-Scl和Ku，以获得一种免疫原性抗原。 诊断探针的包容性和代表性面板， 包括这种疾病的临床变异。

A characteristic feature of many rheumatic diseases is the presence of circulating antinuclear antibodies. Assays based on the measurement of these antibodies are established in the clinical laboratory for the diagnosis of these diseases, but often without the benefit of having well characterized or isolated target antigens. One such autoimmune disease is scleroderma or progressive systemic sclerosis. The objective of this proposal is to provide sensitive and quantitative diagnostic/prognostic assays for this disease by developing assays based on the direct binding of serum antibodies to recombinant polypeptide antigens. The initial focus will be on the five polypeptides identified with the two major scleroderma-associated antigens, scl-70 and centromere. The antigens will be isolated in quantities sufficient for microsequencing. Oligonucleotide probes based on these sequences will be used to clone the genes coding for these polypeptides from human cDNA libraries. Expression of the cloned genes will allow for the synthesis of gram amounts of antigen sufficient for the development of the assay. The recombinant polypeptides will also be used as immunogens to insure a constant and abundant supply of antibodies for antigen isolation and ultimately assay standardization. Similar efforts will be directed toward other of the scleroderma-associated antigens, including fibrillarin, PM-Scl and Ku, to obtain an inclusive and representative panel of diagnostic probes to encompass the clinical variants of this disease.