功能化PMO微球的设计制备及其在分化抗原簇糖蛋白定量检测分析中的应用

具有临床诊断价值的分子标志物往往以低丰度蛋白、多肽的形式表达于生物体系中，其定量检测在生理疾病诊断及分子生物病因探寻上发挥着关键性作用，但在实际样品分析中应用难度极大；而更大的难题是发展简单、灵敏、定量的方法以选择性分析分子标志物后翻译修饰作用，比如糖基化。针对以上问题，研究发现使用多孔材料富集蛋白、多肽可以极大提高检测分析效率。课题拟制备一系列具有不同结构参数的功能化周期性介孔有机硅（PMO）材料，并集成于微球器件表面作为富集检测的微装置；通过系统研究其结构参数与分析效率之间的关系，建立一种面向分子标志物、无需同位素标记的定量质谱分析方法。以上技术路线将应用于分化抗原簇蛋白中糖基化CD44（g-CD44）的检测，该蛋白作为一种典型的分子标志物，与肿瘤干细胞密切相关。我们拟在实际样品分析中定量检测g-CD44浓度，与肿瘤的发病转移过程关联，以期在临床诊断中理解肿瘤的发病转移机制。

Biomarkers with clinical significance are most often various forms of proteins/peptides at very low concentrations in biological systems. Their quantitative detection is important to diagnose and understand the molecular basis of disease, but very difficult in bio-samples. A promising method to enhance the detection efficiency is to use nanoporous materials to enrich proteins or peptides. Despite the pioneering work in this field, little has been done to construct smart, tangible devices from the nanomaterials and investigate the influence of their structural parameters on enrichment and detection efficiency of low-abundance bio-molecules. A further challenge is to develop a simple, sensitive and quantitative method to selectively detect post-translational proteins, such as glycosylated proteins. In this project, a series of periodic mesoporous organosilicas (PMO) decorated beads with controlled structures and tuneable surface chemistry will be synthesised. The correlation between their structural parameters and enrichment/detection efficiency for standard peptides and proteins will be comprehensively studied. Beads decorated by optimized PMO will be used to develop a label-free protocol for the quantitative analysis of standard peptides/proteins by mass spectroscopy (MS). The developed protocol will be employed in the detection of glycosylated cluster of differentiation antigen protein (g-CD44), a biomarker hypothesised to be highly associated with cancer stem-like cells (so called CSC) and the development of cancer. The correlation of quantitative g-CD44 levels with cancer developing stages will be analysed in biological samples harvested from animal model and patients. Successful and quantitative analysis of g-CD44 levels has a potential significance in understanding the mechanism, diagnosis and clinical treatments of cancer, a disease that affects millions of people worldwide.