Simultaneous Detection of Multiple Proteins Using Functionalized Metal Nanoparticles Coupled with Microporous Membrane Substrate and Laser Desorption/ionization Mass Spectrometry

• Main Authors: Chang, Chia-Yin, 張家寅
• Other Authors: Huang, Chih-Ching
• Format: Others
• Language: zh-TW
• Published: 2016
• Online Access: http://ndltd.ncl.edu.tw/handle/nxubnh

碩士 === 國立臺灣海洋大學 === 生命科學暨生物科技學系 === 104 === In this work we demonstrate a simple assay for the simultaneous detection of multiple proteins by functionalized nanomaterials assisted laser-induced desorption/ionization mass spectrometry (LDI-MS). We chose thrombin, vascular endothelial growth factor-A165 (VEGF-A165) and platelet-derived growth factor-BB (PDGF-BB) as model proteins to demonstrate our sensing method. Selective detection of these analytes was achieved by modifying nanoparticles of three different metals (Au, Ag and Pt) with their corresponding aptamer or antibody. Functionalized metal nanoparticles (NPs) bound to their specific analyte was subjected to LDI-MS on a cellulose acetate membrane (CAM). The microporous CAM served as a medium for an in situ separation of NPs based on the increase in particle weight and/or decreased affinity towards CAM with increasing analyte concentration. As a result, the intensities of the signals of the metal cluster ions ([Mn]¬¬+; M = Au or Ag n = 13) decreased in the mass spectra under pulsed laser irradiation. Platinum cluster ion signal being the only exception, signal increases with analyte concentration, due to aggregation of Pt NPs in the presence of PDGF-BB. The nanoparticles/CAM system can function as a signal transducer and amplifier through transformation of protein concentration to intense metal clusters ions signal during LDI-MS analysis. Our developed system can selectively detect proteins down to picomolar region. Most importantly, the system is able to detect multiple proteins simultaneously without any pre-concentration, separation or purification process since LDI-MS coupled with CAM effectively removes all signals except metal cluster ions. This sensing system shows great potential for the simultaneous detection of different protein, tumor markers, pathogens, and cancer cells by functionalizing with suitable ligands to nanomaterials.