Introducing specificity and sensitivity in imaging mass spectrometry

在这项研究计划中，我们将利用我们在成像质谱（IMS）方面的专业知识来解决生物成像领域未满足的重要需求。因此，本提案将侧重于开发分析方法，以提高IMS分析几种临床相关内源性分子的特异性和敏感性。必须开发敏感和有针对性的方法，以促进识别可用于疾病诊断或预后或作为治疗反应指标的分子靶标。这种有针对性的方法也可以在法医学领域得到发展。将探索三个主要的研究方向：1)开发化学定制表面，以特别增强对薄组织切片的脂质和代谢物的检测，同时保持其空间完整性。纳米材料的快速发展促进了表面辅助LDI IMS （SALDI IMS）的出现，为低分子量化合物的成像开辟了新的前景。我们建议开发几种基于金和硅的SALDI表面，用不同的化学物质进行精确的织构和处理，以增强解吸/电离和分析物的特异性。此外，更好地控制表面疏水性将大大减少分析物的离域。2)提出了原位衍生化化学的发展，用于组织切片解吸电喷雾电离（DESI） IMS分析多肽、脂质和代谢物。多肽（和蛋白质）是更具挑战性的生物分子分析DESI IMS从组织切片。肽N或c端衍生化使用含有固定电荷的亲水性反应基团的新化学物质增强肽的溶解度并消除电喷雾电离离子抑制效应。这种反应性标签可以在IMS过程之前或期间应用于组织（反应性DESI）。3)将开发创新方法，专门检测和绘制潜在指纹中包含的遗传物质，以确定可疑性别。这方面是法医界最感兴趣的。为了靶向性别特异性基因，互补DNA探针将通过光可切割基团偶联到带电的报告离子上。在定向喷雾沉积和探针培养后，可光切割报告离子在潜在指纹中的精确位置可以通过LDI IMS绘制出来。通过他的研究计划，IMS方法开发的持续努力将允许对法医和生物样品进行更深入和独特的分子分析，补充我们实验室已经开发和可用的分子分析。总的来说，这些新方法将广泛应用于包括人类疾病和失调以及理解细胞、器官和整个生物体的正常发育和功能在内的生物学应用。

In this research proposal, we will leverage our expertise in Imaging Mass Spectrometry (IMS) to address important unmet needs in the field of biological imaging. Accordingly, this proposal will focus on the development of analytical methods to improve the specificity and sensitivity of IMS for the analysis of several classes of clinically relevant endogenous molecules. Sensitive and targeted methods must be developed to facilitate the identification of molecular targets that can be used in the diagnosis or prognosis of disease or as indicators of therapeutic response. Such targeted approaches can also be developed in the field of forensic science. Three main axes of research will be explored: 1) The development of chemically tailored surfaces to specifically enhance the detection of lipids and metabolites from thin tissue sections while maintaining their spatial integrity. The emergence of Surface-Assisted LDI IMS (SALDI IMS), fostered by the rapid development of nanomaterials, has created new prospects for the imaging of low molecular weight compounds. We propose to develop several gold and silicon-based SALDI surfaces precisely textured and treated with various chemistries to enhance desorption/ionization and analyte specificity. Further, a better control of surface hydrophobicity will greatly minimize analyte delocalization. 2) The development of in situ derivatization chemistries for the analysis of peptides, lipids and metabolites by desorption electrospray ionization (DESI) IMS from tissue sections is proposed. Peptides (and proteins) are more challenging biomolecules to analyze by DESI IMS from tissue sections. Peptide N or C-terminal derivatization novel chemistries using hydrophilic reactive groups containing a fix charge enhances peptide solubility and eliminates electrospray ionization ion suppression effects. Such reactive tags can be applied on tissue before or during (reactive DESI) the IMS process. 3) Innovative approaches will be developed to specifically detect and map genetic material contained within latent fingerprints with the goal of identifying the suspect gender. This aspect is of prime interest to the forensic community. To target gender specific genes, complementary DNA probes coupled to a charged reporter ion via a photocleavable group will be developed. After on-target spray deposition and probe incubation, the precise location of the photocleavable reporter ion within the latent fingerprint can then be mapped by LDI IMS. Through his research program, continued efforts in IMS method development will allow a deeper and unique analysis of molecules from forensic and biological samples complementary to those already developed and available in our laboratory. Overall, these new methods will be useful for a wide range and biological applications including human diseases and disorders as well as understanding normal development and functioning of cells, organs and whole organisms.