Bio-Mass Spec Microanalysis augments Optical Histology

描述(申请人提供)：用亚微米聚焦激光解吸质谱仪对单个哺乳动物细胞进行有意义的分子分析一直是一个难以实现的目标。然而，这种微量分析是通过我们独特的MALDI-离子迁移率-正交飞行时间质谱仪(MALDI-IM-OTOFMS)实现的。将特殊纳米颗粒基质(NP)的亚单分子层应用到亚细胞组织体积上，使微聚焦激光能够解吸中性分析物分子(然后用第二个激光后电离并进行质量分析)。因此，申请者现在的主要目标是创造和优化新的纳米颗粒基质和技术，以便将NPs快速传播到亚细胞体积中。我们将(1)展示通过NP注入的亚细胞分子图谱，并与MALDI组织成像中使用的基质进行比较；(2)设计纳米颗粒，促进中性分析物的非破坏性激光解吸，然后用第二束激光进行后电离；(3)通过重复基质植入循环，然后使用MALDI-IM-OTOFMS(带后电离)进行分子分析，展示细胞的分子深度图谱。这一方法将与DNA、RNA和f-肌动蛋白的几种标准组织学染色(以及其他)进行比较。我们认为这些发展是将定量微量分析方法转变为对基础研究和病理分析有用的亚细胞分子结构的生化成像的关键桥梁技术。 公共卫生相关性：Ionwerks纳米粒子植入器现已上市销售，申请者描述的创新可以快速测试和集成，以增强一种已经很有用的仪器和技术的能力，该仪器和技术还处于商业初级阶段。纳米颗粒基质的快速植入是细胞内分析的关键技术，而细胞内分析是分子表型分析的基础。因此，实现基于细胞水平的质谱学分子表型鉴定似乎是可行的，并将是生物医学科学和临床病理学的重要发展。简单的方法，如质谱仪图像和光(或共焦)显微图像的重叠，应提供快速、简化、广泛的分子表型，即使是小的或有限的活检样本。

DESCRIPTION (provided by applicant): Meaningful molecular analysis of single mammalian cells by submicron focused laser desorption mass spectrometry has been an elusive goal. However, such microanalysis has been achieved with our unique MALDI-Ion Mobility-orthogonal Time of Flight Mass Spectrometry (MALDI-IM-oTOFMS). Applying submonolayers of special nanoparticulate matrices (NP) onto sub cellular tissue volumes enables a micro focused laser to desorb neutral analyte molecules (which are then post-ionized with a second laser and mass analyzed). Thus, the applicants' main goal now is the creation and optimization of new nanoparticulate matrices and techniques for rapid dissemination of NPs into sub cellular volumes. We will (1) demonstrate sub cellular molecular profiling by NP implantation and compare against matrices used in MALDI tissue imaging (2) design nanoparticulates which promote the non-destructive laser desorption of neutral analytes which can then be post-ionized with a second laser, and (3) demonstrate molecular depth profiling of cells through repeated cycles of matrix implantation followed by molecular analysis by MALDI-IM-oTOFMS (with post- ionization). This approach will be compared with several standard histological stains for DNA, RNA and f-actin (among others). We consider these developments to be critical bridging technologies for transforming quantitative micro-analytical methods into biochemical imaging of sub cellular molecular structure useful for basic research and pathological analysis. PUBLIC HEALTH RELEVANCE: The Ionwerks nanoparticulate implanter is now offered for sale, and the innovations described by the applicants can be rapidly tested and integrated to augment the power of an already useful instrument and technique which is in its commercial infancy. The rapid implantation of nanoparticulate matrices is essential technology for intracellular analysis and intracellular analysis is the basis for molecular phenotyping. Thus achieving cellular level mass spectrometry-based molecular phenotyping seems feasible and would be an important development in biomedical sciences and clinical pathology. Simple approaches, such as overlays of the mass spec image and light (or confocal) microscopic images should provide fast, streamlined, broad molecular phenotyping of even small or limited biopsy samples.