RADIOLABELING OF FABS BY PHOTOCHEMICAL CONJUGATION

描述：拟议的研究是为了准备和评估双功能 用于光标记抗体和 与放射性核结合的抗体片段(Te-99m和Re-186/Re-188)。这些 研究是基于一项重大发现，即光标记 蛋白质和抗体可以用全氟芳基叠氮光探针 以1：1的比例实现了高效率(75-80%)的光电探针： 抗体，并且几乎完全保留了抗体的免疫反应性 抗体。这些性质是放射性核标记的理想选择，并建议 光标记技术在抗体标记方面具有真正的潜力 治疗性和诊断性放射性核。这项拟议的研究有几个 目的：合成放射性标记的光探针；附着于 抗体和抗体片段；以及体外和体内检测 稳定性、免疫反应性、生物定位和清除。首字母 努力将不能发光的部分连接到特定的核上 已知具有良好稳定性的螯合剂。依恋将会 通过使用光电探针和螯合剂之间的连接物来实现 确保电子和空间隔离。链接器也将被设计 以促进机体分解代谢后的放射性同位素清除。 这些合成的双功能螯合剂将被标记为 用于测试稳定性和光标记效率的所需放射性金属。 这些用于抗体标记的双功能放射性标记的可行性将 用B72.3抗体和限量生产的抗体片段进行检测 在细菌中进行蛋白分解或重组表达。贴标签效率将 通过使用紫外线和/或辐射监测器的尺寸排除高效液相色谱进行测定 使用先前开发的方法。那些显示出高度依附性的标签 效率将进一步评估使用体外稳定性研究和 免疫反应性。最后，标签材料，它显示了最伟大的 承诺，将接受体内生物分布测试和 净空

DESCRIPTION: The proposed research is to prepare and evaluate bifunctional chelating agents designed for the purpose of photolabeling antibodies and antibody fragments with radionuclei (Tc-99m and Re-186/Re-188). These studies are based on the significant discovery that the photolabeling of proteins and antibodies with a perfluoroaryl azide photoprobe could be achieved with high efficiency (75-80%) in a 1:1 ratio of photoprobe: antibody, and with nearly complete retention of immunoreactivity of the antibody. These properties are ideal for radionuclear labeling and suggest that photolabeling has real potential for labeling of antibodies with therapeutic and diagnostic radionuclei. The proposed research has several objectives: synthesis of the radiolabeled photoprobe; attachment to antibodies and antibody fragments; and in vitro and in vivo testing of stability, immunoreactivity, biolocalization, and clearance. The initial effort will be the attachment of the photolabile moiety to nuclei-specific chelating agents, which are known to have good stability. Attachment will be achieved using linkers between the photoprobe an the chelating agents to insure electronic and spatial isolation. The linkers will also be designed to promote radioisotope clearance following catabolism b the organism. These synthesized bifunctional chelating agents will be labeled with the desired radiometal for testing of stability and photolabeling efficiency. The viability of these bifunctional radiolabels for antibody labeling will be tested with the B72.3 antibody and antibody fragments produce by limited proteolysis or recombinant expression in bacteria. Labeling efficiency will be determined by size exclusion HPLC using UV and/or radiation monitors using methods developed earlier. Those labels showing high attachment efficiency will be further evaluated using in vitro studies of stability and immunoreactivity. Finally, the labeled materials, which show the greatest promise, will be subjected to in vivo testing for biodistributon and clearance