High-spatial-resolution mass spectrometry imaging of biological tissues using a microfluidic probe

Nanospray desorption electrospray ionization (nano-DESI) is a liquid extraction-based ambient ionization mass spectrometry imaging (MSI) technique that enables quantitative molecular mapping of biological samples in their native state with high spatial resolution. To facilitate the wider adoption of nano-DESI MSI by the scientific community, we have developed a robust and user-friendly microfluidic probe (MFP). The probe has been used to achieve high spatial resolution of 8-10 µm and up to 10-fold improvement in the experimental throughput, enabling imaging of large tissue sections with cellular resolution. Here, we provide detailed instructions for designing, fabricating and operating MFPs. In addition, we describe a complete workflow for nano-DESI MSI, covering every step from probe assembly to data acquisition and analysis. Although the fabrication of MFPs requires expertise in microfluidics and can take a few days, the process can be outsourced to qualified companies for manufacturing. Once the MFP is fabricated, the entire imaging workflow can be completed in several hours, depending on the sample size. For example, a sample with an area of 1 cm² can be analyzed in <10 h at a spatial resolution of 10 µm. The exceptional performance and ease of use of these probes will make high-resolution nano-DESI MSI more accessible to the scientific community.