CHARACTERIZATION OF THE ANTI-RH ALLOIMMUNE RESPONSE

通过输血或输血对猕猴血型系统进行同种异体免疫 怀孕会限制输血的有效性，可能会导致 临床上有显著的输血反应，并可引起溶血 新生儿疾病。为了设计出具体有效的方法 用于预防和管理与Rh有关的问题 同种异体免疫，首先要鉴定抗-Rh免疫 在分子水平上的反应。 为此，拟议的研究将利用一些新的细胞 和重组方法来检查可变区的谱系 抗Rh同种异体抗体使用的基因以及结构 它们的抗原结合部位的异质性。具体来说，我们将 人抗Rh同种异体抗体在人B细胞中的体外表达 首先用交联型抗-HBs扩增B细胞，使患者致敏 CD40和IL-4，然后在筛选产生抗Rh的克隆之前 EBV或体细胞杂交转化细胞。同时， 丝状噬菌体免疫球蛋白展示文库将从 B细胞的初始群体，以及从扩增和转化 克隆，以挽救不稳定或难以永生的细胞。 这种结合的方法将提供表达的免疫球蛋白和 编码它们的DNA。这将使得能够识别 Rh抗原的免疫优势表位和抗-HBs的保守DNA序列 RH抗体。最后，我们将分离抑制抗Rh抗体的多肽 通过分析高变环与红细胞的体外结合 封闭(抗独特型)抗体及随机多肽的筛选 具有抗Rh抗体的文库。 总而言之，这些研究将提供对 在分子遗传水平上人类对Rh抗原的免疫反应。 这将促进设计治疗上有用的抑制剂， 调节体内抗Rh/Rh抗原的相互作用。它还将导致 调节抗Rh/Rh抗原的基因疫苗的研制 活体内的相互作用。它还将导致基因的发展 诱导T细胞反应的疫苗可下调aA的产生 抗Rh同种异体抗体。此外，体外生产的单抗 抗Rh抗体/重组Fab片段及其结构类似物 它们的抗原表位将提供有用的生化和血清学 增加我们对生物化学和免疫学的了解的试剂 这个重要的血型系统。

Alloimmunization to the Rhesus blood group system through transfusion or pregnancy can limit the effectiveness of transfusions, can lead to clinically-significant transfusion reactions, and can cause hemolytic disease of the newborn. In order to design specific and effective methods for the prevention and management of the problems associated with Rh alloimmunization, it is first necessary to characterize the anti-Rh immune response at the molecular level. To this end, the proposed research will utilize a number of novel cellular and recombinant approaches to examine the repertoire of variable regions genes used by anti-Rh alloantibodies as well as the structural heterogeneity of their antigenic binding sites. Specifically, we will express human anti-Rh alloantibodies in vitro from the B-cells of sensitized patients by first expanding the B-cells with crosslinked anti- CD40 and IL-4, and then screening for anti-Rh producing clones prior to cell transformation with EBV or somatic cell hybridization. In parallel, filamentous phage immunoglobulin display libraries will be created from the initial populations of B-cells, as well as from expanded and transformed clones, to rescue cells that are unstable or refractory to immortilization. This combined approach will provide both the expressed immunoglobulins and the cDNAs that encode them. This will enable the identification of immunodominant epitopes on Rh antigens and conserved DNA sequences on anti- Rh antibodies. Finally, we will isolate peptides that inhibit anti-Rh binding to RBCs in vitro through the analysis of hypervariable loops of blocking (anti-idiotypic) antibodies and the screening of random peptide libraries with anti-Rh antibodies. Collectively, these studies will provide an understanding of the nature of the human immune response to Rh antigens on a molecular genetic level. This will facilitate t he design of therapeutically useful inhibitors that modulate anti-Rh/Rh antigen interactions in vivo. It will also lead to the development of genetic vaccines that modulate anti-Rh/Rh antigen interactions in vivo. It will also lead to the development of genetic vaccines that induce T-cell responses that down-regulate athe production of anti-Rh alloantibodies. In addition, the in vitro production of monoclonal anti-Rh antibodies/recombinant Fab fragments and the structural analogs of their antigenic epitopes will provide useful biochemical and serological reagents to increase our understanding of the biochemistry and immunology of this important blood group system.