In Situ Tissue Analysis of Neuropeptides by MALDI FTMS In-Cell Accumulation

Abstract

Herein we report the first application of Fourier transform mass spectrometry for the analysis of neuropeptides directly from neuronal tissues. Sample preparation protocols and instrumentation conditions are developed to allow in situ neuropeptide analysis of the neuroendocrine organs freshly isolated from a marine organism Cancer borealis. The utility of a previously developed in-cell accumulation (ICA) technique is extended for peptide analysis in complex tissue samples. With the ICA procedure, ion signals from multiple laser shots are accumulated in the analyzer cell prior to detection. This procedure allows the accumulation of ion signals without accumulating noise, thus improving the signal-to-noise ratio and enhancing the sensitivity for the detection of trace-level endogenous neuropeptides. De novo sequencing of peptides directly from tissue samples becomes more feasible through this improvement. Additionally, an integrated pulse sequence is constructed to cover a wide mass range from m/z 215 to 9000 by centering quadrupole collection of ions at different masses for successive laser shots. Finally, improved mass measurement accuracy (2 ppm) for tissue peptide analysis is achieved using ICA by incorporating calibrants on a separate spot from the sample of interest without premixing calibration standards with the analytes.

Identification of Glioma Neovascularization-related Proteins by Using MALDI-FTMS and Nano-LC Fractionation to Microdissected Tumor Vessels

“…Recent studies showed that the application of MALDI-FTMS holds significant advantages over other types of mass spectrometry (21,22). FTMS provides very high mass accuracy, which is considerably increased by its ability to perform an internal calibration (23).…”

Midgut epithelial endocrine cells are a rich source of the neuropeptides APSGFLGMRamide (Cancer borealis tachykinin-related peptide Ia) and GYRKPPFNGSIFamide (Gly1-SIFamide) in the crabs Cancer borealis, Cancer magister and Cancer productus

“…Desalted extracts were mixed 1:1 with DHB matrix (150·mg·ml -1 in 50% methanol/deionized water) on a MALDI probe tip and allowed to crystallize at room temperature. MALDI-FTMS analysis was then performed as described in several recent publications (Kutz et al, 2004;Messinger et al, 2005) using an IonSpec HiResMALDI Fourier transform mass spectrometer equipped with a 7.0-T actively shielded superconducting magnet (School of Pharmacy, University of Wisconsin-Madison).…”

Imaging of peptides in the rat brain using MALDI-FTICR mass spectrometry

“…By using MALDI-FTICR (Fourier transform ion cyclotron resonance) MS it is possible to combine high resolving power, high mass accuracy, and the possibility of performing multistage MS/MS experiments [35][36][37][38] allows peptide identification directly from tissue [39].°The°FTICR°imaging°strategy°extends°the°identifi-cation possibilities in direct tissue analysis and is a valuable tool in MS imaging instrumentation.…”