Development of VSSI-probe technology for in situ probing biological systems using mass spectrometry

项目摘要 质谱是生物学研究的有力工具，具有广泛的应用前景 包括蛋白质组学、代谢组学、糖组学、脂质组学和结构生物学。虽然大多数现有的MS 应用是基于提取的批量分析，使用MS对生物样品进行原位或体内分析 对于新的生物学发现变得越来越重要。过去十年见证了 快速开发用于直接MS分析来自组织切片的各种生物样品的接口方法 单个细胞。然而，现有的方法在研究活的和动态的生物系统时仍然存在困难 在一个非侵入性的方式和原位MS实验主要是由MS研究小组进行，由于 涉及复杂的仪器设置。因此，这些限制严重阻碍了使用 直接MS用于生物学发现。为了解决生物系统的原位MS分析的未满足的需求， 我们的目标是发展一种直接质谱方法，振动锐边喷雾电离探针（VSSI-probe）， 是简单的、易于使用的、生物相容的，并且适合于高速采样。与现有方法相比， 所提出的VSI探针适于快速采样、高度生物相容、无电压、易于操作，并且 便宜得多。因此，我们预计，一旦证明，拟议的VSSI-probe将使 新的MS应用程序，以及提高直接MS分析在生物界的采用。在 具体而言，我们将（1）优化用于高灵敏度MS检测的VSI探针的性能;（2） 展示用于研究快速生物反应的高通量VSI探针平台;（3）展示 VSSI-probe在研究脂质/蛋白质聚集体相互作用的微妙和动态生物系统中的实用性。 (4)展示了用于直接研究代谢物的VSI探针的高空间分辨率采样能力 细菌菌落。凭借探测生物样品的前所未有的能力和灵活性， 所提出的VSSI-probe将使广泛的生物应用成为可能， 采样包括酶学、细胞监测、脂质双层化学和真实的实时术中诊断。 所提出的系统的简单性、低成本以及与现有质量分析仪的兼容性也将大大提高系统的性能。 减少非MS群体采用该技术的技术障碍，从而扩大 将MS导向更多样化的生物学问题。

Project Summary Mass spectrometry (MS) is a powerful tool for biological studies, which has found a wide range of applications including proteomics, metabolomics, glycomics, lipidomics, and structural biology. While most of the existing MS applications are extraction-based batch analysis, using MS for in situ or in vivo analysis of biological samples has become increasingly important to enable new biological discoveries. The past decade has witnessed the rapid development of interfacing methods for direct MS analysis of various biological samples from tissue slices to single cells. However, existing methods still have difficulties in studying living and dynamic biological systems in a non-invasive manner and in situ MS experiments are primarily performed by MS research groups due to the involvement of complex instrument setups. As a result, these limitations significantly hinder the progress of using direct MS for biological discoveries. To address the unmet need for in situ MS analysis of biological systems, our objective is to develop a direct MS method, vibrating sharp-edge spray ionization probe (VSSI-probe), that is simple, easy to use, biocompatible, and amenable to high speed sampling. Compared to existing methods, the proposed VSSI-probe is amenable to fast sampling, highly biocompatible, voltage-free, easy to operate, and substantially less expensive. Therefore, we expect that once demonstrated, the proposed VSSI-probe will enable new MS applications as well as improve the adoption of direct MS analysis in the biological community. In particular, we will (1) optimize the performance of the VSSI-probe for high sensitivity MS detection;(2) demonstrate a high-throughput VSSI-probe platform for studying fast biological reactions;(3) demonstrate the utility of VSSI-probe in studying a delicate and dynamic biological system of lipid/protein aggregates interaction. (4) demonstrate the high spatial resolution sampling capability of VSSI-probe for studying metabolites directly from bacterial colony. With the unprecedented capability and flexibility of probing biological samples, the proposed VSSI-probe will enable a broad range of biological applications that requires fast and biocompatible sampling including enzymology, cell monitoring, lipid bilayer chemistry, and real time intraoperative diagnosis. The simplicity, low cost, and compatibility with existing mass analyzers of the proposed system will also greatly reduce the technical barrier for adopting this technology by non-MS groups thereby expanding applications of direct MS to more diverse biological problems.