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Abstract Nanospray desorption electrospray ionization (nano‐DESI) is an ambient ionization mass spectrometry imaging (MSI) approach that enables spatial mapping of biological and environmental samples with high spatial resolution and throughput. Because nano‐DESI has not yet been commercialized, researchers develop their own sources and interface them with different commercial mass spectrometers. Previously, several protocols focusing on the fabrication of nano‐DESI probes have been reported. In this tutorial, we discuss different hardware requirements for coupling the nano‐DESI source to commercial mass spectrometers, such as the safety interlock, inlet extension, and contact closure. In addition, we describe the structure of our custom software for controlling the nano‐DESI MSI platform and provide detailed instructions for its usage. With this tutorial, interested researchers should be able to implement nano‐DESI experiments in their labs.more » « less
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