A new method to improve sensitivity and resolution in matrix‐assisted laser desorption/ionization time of flight mass spectrometry

Abstract

High detection sensitivity and resolution are two critical parameters for recording good peptide mass fingerprints (PMF) of low abundance proteins. This paper reports a mass spectrometry (MS) sample preparation technique that could improve sensitivity and resolution. By coating the MS steel target with a thin layer of pentadecafluorooctamido propyltrimethoxysilane, which was both polar and nonpolar solvent repellent, the transferred sample droplets on its surface were significantly smaller. As a result, the analyte of the peptide mixture became more concentrated and homogeneous, which helped to improve the sensitivity. The advantages of a modified MS target were documented by mass spectra improvement of attomole level standard peptides and silver-stained proteins from polyacrylamide gels. The mass signal of angiotensin II at 100 attomole was difficult to record on the conventional support, whereas it was easily detected on the modified one. The PMF of cytochrome C was also better recorded on the modified support, in terms of both signal-to-noise ratio and the number of detected peptides. When silver-stained proteins from two-dimensional electrophoresis gels were analyzed, in most cases more satisfactory peptide mass spectra were obtained from the modified support. Searching protein databases with more mass data from the improved PMFs, several unknown proteins were successfully identified.

Proteomic Analysis of Low Nitrogen Stress-Responsive Proteins in Roots of Rice

“…MS analysis was carried out for 21 differentially expressed proteins resulting in 12 proteins with a significant match, while the rest showed no significant match in the database. MS is typically able to detect only a few peptide mass peaks for low abundance protein spots (Xiong et al 2003). We were able to detect more spots, but for these low abundance proteins, database searching becomes difficult and the results become less credible.…”

Matrix‐assisted laser desorption/ionization of peptides on AnchorChip™ targets with α‐cyano‐4‐hydroxycinnamic acid and nitrocellulose as matrix

“…Gobom et al [7][8][9][10][11] or sample supports with hydrophobic surfaces, [9][10][11][12][13] have been reported by several groups over the last few years. The concentration of CHCA and the composition of the solvents in these reports vary considerably.…”

Interfacing microfluidics to LDI-MS by automatic robotic spotting

“…Several microfabricated MALDI target plates have been demonstrated to physically constrain lateral sample spreading; these modifications have included anisotropically bulk etching on silicon target plate to create vials (Ekstrom et al 2001a;Little et al 1997) or directly milling to create vials on polymer-based target plate to increasing local concentration (Ekstrom et al 2001b). The most common approach is done by either coating hydrophobic layer as sample support like poly(tetrafluoroethylene) (Hung et al 1999) and propyltrimethoxysilane (Xiong et al 2003) or by coating hydrophilic gold pad on hydrophobic Teflon background (Schuerenbeg et al 2000) to constrain lateral spreading of analyte during deposition and matrix cocrystallization. In a recent research demonstration, galvanic etching techniques have also been used to produce nanoscale porous or filament-like structure on silicon surfaces as matrix-free, high sensitivity LDI-MS target substrate for bioanalysis.…”