Optimization of the use of Py–Tag for next generation derivatization reagents in imaging mass spectrometry

Abstract

To demonstrate the accurate analysis of catecholamines and amino acid using derivatization reagents, we investigated the reaction conditions for 2,4,6-triethyl-3,5-dimethyl pyrylium trifluoromethanesulfonate (Py-Tag), derivatization of the targets dopamine (DA) and γ-aminobutyric acid (GABA) on tissue sections, and constructed an optimized reaction compartment. Ten different Py-Tag reaction conditions with the targets were considered. The optimal condition for the Py-Tag reaction with the targets was identified as a 70% methanol with 5% trimethylamine (v/v) solution at 60°C under homogenous conditions. To reproduce this reaction on tissue sections, we constructed a reaction compartment to maintain humidity levels and facilitate the derivatization reaction. Moreover, visualization of DA and GABA was archived by derivatized-imaging mass spectrometry. Brain sections of unilateral 6-OHDA lesioned Parkinson's disease model rats showed Py-Tag DA (m/z 328.3) in the unilateral striatum and Py-Tag GABA (m/z 278.3) in the cerebral cortex, striatum, hippocampus and hypothalamus. Using the Parkinson's disease model rat brain, images with left-right differences were obtained for the localization of DA and GABA. These findings indicate that it is important to consider the reaction conditions that allow high reaction efficiency between DA or GABA and Py-Tag as well as high quality imaging of sections.