Automated Spatially Targeted Optical Micro Proteomics (AutoSTOMP) 2.0 identifies proteins enriched within inflammatory lesions in tissue sections and human clinical biopsies

Abstract

ABSTRACTTissue microenvironment properties like blood flow, extracellular matrix or proximity to immune infiltrate are important regulators of cell biology. However, methods to study regional protein expression in context of the native tissue environment are limited. To address this need we have developed a novel approach to visualize, purify and measure proteins in situ using Automated Spatially Targeted Optical Micro Proteomics (AutoSTOMP) 2.0. We previously implemented AutoSTOMP to identify proteins localized to the vacuoles of obligate intracellular microbes at the 1-2 μm scale within infected host cells1. Here we report custom codes in SikuliX to specify regions of heterogeneity in a tissue section and then biotin tag and identify proteins belonging to specific cell types or structures within those regions. To enrich biotinylated targets from fixed tissue samples we developed a biochemical protocol compatible with LC-MS. These tools were applied to a) identify inflammatory proteins expressed by CD68+ macrophages in rat cardiac infarcts and b) characterize inflammatory proteins enriched in IgG4+ lesions in esophageal tissue. These data indicate that AutoSTOMP is a flexible approach to determine regional protein expression in situ on a range of primary tissues and clinical biopsies where current tools are limited.