SBIR Phase I: In vivo Fluorescence Imaging Kit for Cell Proliferation.

这个小企业创新研究（SBIR）一期项目是将新型生物正交四嗪连接技术应用于DNA复制的荧光标记，并最终生产用于细胞增殖的荧光成像试剂盒（FIK）。其优势有两方面：1)由于新的化学机制，FIK可以在体内进行细胞成像，基本上可以在细胞代谢干扰最小的情况下连续观察细胞增殖。荧光显微镜可用于活细胞。2)单个细胞可以通过细胞周期的各个阶段进行监测。现有的方法都不允许对细胞增殖进行体内成像。因此，从概念上讲，现有测定法和提议的FIK测定法的能力之间的差异类似于快照和视频记录之间的差异。该项目的更广泛的影响/商业潜力是开发一种与高通量筛选技术兼容的细胞增殖试验。该试验可用于区分静止和活跃循环的干细胞，并最终用于区分癌症干细胞和癌细胞。试剂成本低，样品处理简单，易于适应多孔格式，使所提出的方法成为高通量筛选分析的理想候选者，将在癌症生物学和神经发生，以及发育和分子生物学领域找到应用。这些领域都有一个共同的研究需求，即密切观察细胞过程。

This Small Business Innovation Research (SBIR) Phase I project is to apply novel bio-orthogonal tetrazine ligation technology to fluorescent labeling of DNA replication, and to ultimately produce a fluorescence imaging kit (FIK) for cell proliferation. The advantages are two-fold: 1) Because of the novel chemical mechanism, the FIK would allow for in vivo cell imaging, essentially providing continuous observation of cell proliferation with minimal interference from cell metabolism. Fluorescent microscopy could be done with live cells. 2) A single cell could be monitored through various stages of cell cycle. None of the existing methods allow for in vivo imaging of cell proliferation. Thus, conceptually, the difference between the capabilities of the existing assays and proposed FIK assay is analogous to the difference between a snapshot and a video recording. The broader impact/commercial potential of this project is to develop an assay for cell proliferation compatible with high-throughput screening technology. The assay can be applied to differentiate between quiescent and actively cycling stem cells, and, eventually, to differentiate between cancer stem cells and cancer cells. The low cost of reagents, simplicity of sample handling, and readiness to be adapted to a multi-well format make the proposed methodology an ideal candidate for application to high-throughput screening assays that will find applications in the fields of cancer biology and neurogenesis, as well as developmental and molecular biology. These fields all share a common research need for close observation of cell processes.