High sensitivity reagents for bioassays employing mass cytometry

该提案描述了无机纳米颗粒的合成、表征和生物功能化，该纳米颗粒被设计为高灵敏度的细胞计数（CyTOF）试剂，这是一项由我们的合作伙伴Fluidigm加拿大公司发明并商业化的技术。大规模细胞术的工作原理是原子质谱法，并使用标记有金属同位素的试剂来检测单个细胞内或细胞上表达的蛋白质和其他生物标志物。它使人们能够以高通量（高达1000个细胞/秒）检查单个细胞，其最大的优势在于，与使用荧光试剂的流式细胞术相比，它可以同时测量每个细胞更多的生物标志物。目前的一个缺点是它的灵敏度有限，在检测生物标志物存在于低拷贝数每个细胞。这里提出的研究将通过开发纳米粒子试剂来克服这一限制，这种试剂比目前的试剂携带更多的金属同位素拷贝。我们预计纳米颗粒试剂将提供一到两个数量级的灵敏度提高。这项研究的一个主要挑战将是修改这些纳米颗粒的表面，使它们只与靶细胞结合，而与非靶细胞的非特异性相互作用可以忽略不计。此外，我们将开发纳米颗粒试剂，以增加可用于分析的金属同位素的数量，从而增加可用于每个细胞的不同生物标志物的数量。我们希望这些试剂与细胞计数术相结合将提供一种技术，人们可以用来理解、诊断，并有望治愈复杂的人类疾病，如癌症、代谢紊乱和心脏病。

This proposal describes the synthesis, characterization, and biofunctionalization of inorganic nanoparticles designed as high-sensitivity reagents for mass cytometry (CyTOF), a technique invented and commercialized by our partner Fluidigm Canada in Markham ON. Mass cytometry works on the principle of atomic mass spectrometry and employs reagents labeled with metal isotopes to detect proteins and other biomarkers expressed in or on individual cells. It enables one to examine individual cells at high throughput (up to 1000 cells/sec), and its greatest strength is that it can simultaneously measure many more biomarkers per cell than flow cytometry, which employs fluorescent reagents. A current shortcoming of mass cytometry is its limited sensitivity in detecting biomarkers present at a low copy number per cell. The research proposed here will overcome this limitation by developing nanoparticle reagents that carry many more copies of a metal isotope than the current reagents. We anticipate that nanoparticle reagents will provide one to two orders of magnitude increase in sensitivity. A major challenge in this research will be to modify the surface of these nanoparticles so that they only bind to targeted cells and have negligible non-specific interaction with non-targeted cells. In addition, we will develop nanoparticle reagents that will increase the number of metal isotopes that can be employed in an analysis and thus increase the number of different biomarkers that can be measured for each cell. We hope that these reagents in combination with mass cytometry will provide technology that one can use to understand, diagnose, and, hopefully, cure complex human diseases such as cancer, metabolic disorders, and heart disease.