Easy isolation of F4/80 positive macrophages from mouse tissues

Abstract Macrophages are important and also have critical roles in the development and homeostasis of tissues and organs. Our understanding of macrophages is constantly evolving and expanding, as they are actively studied in many areas of research, including infectious diseases, wound healing, and tumor immunology. Adding to the complexity of macrophage research, subsets of macrophages can be identified throughout the body with diverse phenotypes and functions. Furthermore, macrophage frequency can be highly variable across different tissues, presenting a challenge to obtain highly pure macrophages. To address these challenges, we have developed a simple method to isolate macrophages by targeting F4/80, a well-established murine macrophage marker. Starting with a single-cell suspension from mouse peritoneal cavity, lung, or spleen, F4/80-positive cells were labeled with an antibody complex that links F4/80-positive cells to magnetic particles, then separated using an EasySep™ magnet. Using this method, F4/80-positive cells were enriched from 37.3 ± 9.3% to 94.4 ± 2.9% (mean ± SD; n = 12) from peritoneal lavage fluids, 26.5 ± 2.7% to 94.3 ± 2.8 % (n = 9) from lungs, and 8.0 ± 2.4% to 88.8 ± 3.4% (n = 18) from spleens. Protocols have been optimized to accommodate a range of sample sizes from the various tissue sources. EasySep™-isolated F4/80-positive macrophages are functional, as demonstrated by their ability to uptake FITC-dextran and secrete inflammatory mediators upon activation. Overall, the EasySep™ Mouse F4/80 Positive Selection Kit enables simple and easy isolation of F4/80-positive macrophages in under 25 minutes, streamlining the experimental workflows for researchers studying macrophage biology.