High Throughput Biomarker Verification by MALDI-MS Phase II

建立了一种新型的高通量多重亲和捕获平台，用于检测体液中低丰度的肿瘤相关蛋白。该方法利用Labcell Inc.的声波液滴喷射(ADE)技术、生物芯片制造的最新进展、自组装单层表面化学以及基质辅助激光解吸电离质谱仪(MALDI-MS)和其他表面采样质谱学方法，创建了一个具有重复性、灵敏度和动态范围的全自动、多路、亲和捕获平台，具有常规绝对定量低丰度癌症相关蛋白质所需的重复性、灵敏度和动态范围。特别是，该平台将提供比液质联用(LC MS/MS)方法更大的吞吐量，例如使用稳定同位素标准和抗肽抗体捕获的多肽和蛋白质的质谱定量方法(通常称为SISCAPA过程)，同时利用该技术的许多更有吸引力的方面，包括稳定同位素标准和抗肽抗体。随着第二阶段的完成，当与配备5 kHz激光的最先进的MALDI质谱仪一起使用时，开发的自动化平台将能够以至少10倍于基于多反应监测(MRM)的LC-MS/MS方法的速度对96个样品中的蛋白质生物标记物进行多路定量，并且具有类似的灵敏度、重复性和动态范围。

Development of a novel high throughput multiplex affinity capture platform to detect low-abundance cancer related proteins from bodily fluids is proposed. The approach leverages Acoustic Droplet Ejection (ADE) technology from Labcyte Inc., recent developments in biochip fabrication, self-assembled monolayer surface chemistries, and Matrix Assisted Laser Desorption Ionization¿Mass Spectrometry (MALDI-MS) and other surface sampling mass spectrometric approaches to create a fully automated, multiplex, affinity capture platform with the reproducibility, sensitivity, and dynamic range required for routine, absolute quantitation of low-abundance cancer-related proteins. In particular, the platform will offer significantly greater throughput than Liquid Chromatography¿Mass Spectrometry (LC MS/MS) approaches such as the mass spectrometric quantitation of peptides and proteins using stable isotope standards and capture by anti-peptide antibodies (commonly called the SISCAPA process), while exploiting many of the more attractive aspects of that technique including stable isotope standards and anti-peptide antibodies. With completion of Phase II, the automated platform developed when employed in conjunction with a state-of-the-art MALDI mass spectrometer equipped with a 5 kHz laser will enable multiplex quantitation of protein biomarkers from 96 samples at a rate that is at least 10-fold faster than LC-MS/MS approaches based on Multiple Reaction Monitoring (MRM) and with comparable sensitivity, reproducibility, and dynamic range.