Spray-Based Application of Matrix to Agar-Based Microbial Samples for Reproducible Sample Adherence in MALDI MSI

Microbial mass spectrometry imaging (MSI) is a powerful tool used to generate biological hypotheses about the roles of metabolites in microbial competition based upon their two-dimensional spatial distribution. The most commonly used ionization method for microbial MSI is matrix-assisted laser desorption ionization (MALDI). However, medium components and microbial metabolites influence the adhesion of agar samples to the MALDI target, limiting the applicability of MALDI MSI to microbes grown on specific media. Here, we describe a three-step process using a robotic sprayer for a matrix application that improves the adherence of agar samples to the MALDI target, enabling the use of different media for microbial growth and an MSI analysis of larger sample surface areas.