OPTIMIZING TRACERS FOR MULTICOLOR NEURONAL PROFILING

描述（由申请人提供）：PTI Research开发了用于生物医学应用的新型荧光探针。初步数据表明，PTIR 271，一种与共聚焦显微镜的Kr-Ar激光线的647 nm线很好匹配的荧光染料，与其他市售染料相比，具有上级扩散和成像特性，用于固定组织中的神经元分析。与另外两种市售染料PKH 2和PKH 26（其分别由于缓慢的扩散速率和次优的激发而对于神经元分析是次优的）组合，已经证明了神经元分析的三重标记的潜力，但是需要改进。 在SBIR第一阶段，我们将标准化固定组织中的测试系统，用于评估新染料，合成一系列PTIR 271类似物，以了解在固定组织中> 2 mm/hr的有用扩散速率所需的分子结构特征，并通过开发吸光度与Kr-271的所有主要激发线相匹配的化合物来扩展用于神经元成像研究的染料数量。共焦显微镜中的氩激光。SBIR第一阶段的具体目标是：1）标准化测试系统，以评估新染料和完整的评估PTIR 271（Creighton University）; 2）合成PTIR 271类似物以开发SAR信息（PTIR）; 3）至6）产生分别在568 nm、488 nm、752 nm和514 nm处吸收的染料，并且适用于脑神经元成像研究（PTIR）; 7）创建具有> 50 nm的斯托克斯位移的染料，其适用于神经元成像研究（PTIR）;以及8）创建测试试剂盒以将每种染料销售给研究团体（PTIR）。如果可以鉴定一组至少3种荧光亲脂性染料，则证明其可行性，其中： . 该组所有成员的扩散速率不小于2 mm/24小时，CV不大于10%; .组内最大和最小相对染料扩散性（RDD）值相差小于或等于25%; .在用于检测其他集合成员的过滤窗口中，没有成员给出S/N >2X背景; .所有组成员的相对跨细胞标记（RTL）值均不大于市售参比染料Dil。

DESCRIPTION (provided by applicant): PTI Research develops novel fluorescent probes for biomedical applications. Preliminary data has shown that PTIR271, a fluorescent dye well matched to the 647nm line of the Kr-Ar laser line of the confocal microscope has superior diffusion and imaging properties for neuronal profiling in fixed tissue compared to other commercially available dyes. In combination with two other commercially available dyes, PKH2 and PKH26, which are suboptimal for neuronal profiling due to slow diffusion rate and suboptimal excitation respectively, the potential for triple labeling of neuronal profiling has been demonstrated but improvements are needed. In SBIR Phase I, we will standardize test systems in fixed tissue to be used to evaluate new dyes, synthesize a series of PTIR271 analogs to obtain an understanding of structural features of the molecule required for useful diffusion rates of > 2mm/hr in fixed tissue and extend the number of dyes useful for neuronal imaging studies by developing compounds with absorbance's matched to all the major excitation lines of the Kr-Ar laser in confocal microscopes. SBIR Phase I specific aims are to: 1) Standardize test systems for evaluation of new dyes and complete evaluation of PTIR271 (Creighton University); 2) Synthesize PTIR271 analogs to develop SAR information (PTIR); 3) to 6) Create dyes which absorb at 568nm, 488nm, 752nm and 514nm respectively, and are suitable for multicolor neuronal imaging studies (PTIR); 7) Create a dye with a Stokes shift of >50nm which is suitable for multicolor neuronal imaging studies (PTIR); and 8) Create test kits to market each dye to the research community (PTIR). Feasibility will be demonstrated if a set of at least 3 fluorescent lipophilic dyes can be identified for which: . All members of the set diffuse at rates of no less than 2 mm/24 hour with CVs no greater than 10 percent; . Largest and smallest relative dye diffusibility (RDD) values within the set differ by less than or equal to 25 percent; . No member gives S/N >2X background in the filter windows used for detection of other set members; . Relative transcellular labeling (RTL) values for all set members are no greater than the commercial reference dye, Dil.