SBIR Phase I: SERS Enhanced Ligase Detection Reaction Chip for the Molecular Diagnosis of Cancer

这个小型企业创新研究(SBIR)第一阶段项目计划利用新的表面增强拉曼散射(SERS)增强连接酶检测反应(LDR)和光学活性微流控芯片，开发一个全新的癌症分子诊断平台。由于每种染料都有唯一的拉曼指纹，由于光谱重叠较少，可以并行筛选的单核苷酸多态(SNPs)的数量大大增加。第一阶段工作的目标是开发一套目标特定的LDR探头，其格式为6-plex，与当前最先进的格式相匹配。在未来的工作中，我们将开发一种12-复合体格式的分析方法，使用这种基于荧光的技术，可以在一个样本中筛选的靶标数量翻一番。随着筛选反应的发展，我们将创造一种新型的“光流控”芯片，以应对芯片上拉曼信号检测的挑战。该方法的主要优点是通过检测染料的拉曼指纹而不是荧光发射，避免了现有最先进的实时聚合酶链式反应的基本限制。该项目更广泛的影响/商业潜力是能够在不牺牲速度和易用性的情况下筛选更多数量的单核苷酸多态(SNPs)，这可能使癌症和其他遗传疾病的分子诊断更加快速。在某些情况下，SNP是癌症病因中隐含的生物标记物，而在另一些情况下，它们代表癌症风险增加的标记物。在这两种情况下，SNPs已被证明是许多类型癌症的良好生物标记物，并进一步被证明与不同癌症亚型的各种临床病理特征相关。在可用于SNP诊断的众多技术中，实时聚合酶链式反应(RT-PCR)是最适合临床诊断的技术，因为它需要最少的物理样本和样本制备；然而，它在一次筛查的SNP数量方面是有限的。我们提出的方法可能会使单个样本中可以筛查的SNPs数量翻一番。这一优势在活组织检查样本不均一的情况下尤为重要。

This Small Business Innovation Research (SBIR) Phase I project proposes to develop an entirely new platform for the molecular diagnostics of cancer exploiting a new Surface Enhanced Raman Scattering (SERS) enhanced Ligase Detection Reaction (LDR) and an optically active microfluidic chip. Since every dye has a unique Raman fingerprint, the number of single nucleotide polymorphisms (SNPs) that can be screened for in parallel is dramatically increased due to lower spectral overlap. The goal for the Phase I work is to develop a set of target-specific LDR probes in a 6-plex format that would match the current state-of-the-art. For future work, we will develop an assay for a 12-plex format that will double the number of targets that can be screened in one sample using this fluorescence-based technology. In parallel with the development of the screening reaction, we will create a novel "optofluidic" chip that addresses the challenges with on-chip Raman signal detection. The primary advantage of the proposed approach is that it avoids the fundamental limitation of the existing state-of-the-art, Real-Time Polymerase Chain Reaction, by detecting the Raman fingerprint of the dye rather than the florescence emission. The broader impact/commercial potential of this project is the ability to screen for a greater number of single nucleotide polymorphisms (SNPs) without sacrificing speed and ease-of-use, which may enable more rapid molecular diagnosis of cancer and other genetic diseases. In some cases, SNPs are biomarkers implied in the cause of cancer while in others they represent markers indicative of an increased risk of cancer. In either case, SNPs have been shown to be good biomarkers for many classes of cancer and have further been shown to correlate with various clinicopathological features of different cancer subtypes. Of the many technologies available for SNP diagnostics, Real-Time PCR (RT-PCR) is the most appropriate for clinical diagnosis in that it requires the smallest amount of physical sample and sample preparation; however, it is limited in terms of the number of SNPs that can be screened at one time. The approach we propose will potentially double the number of SNPs that can be screened in a single sample. This advantage is particularly important in cases where the biopsied sample is not homogeneous.