Abstract
Fibroblastic reticular cells (FRCs) play critical roles in the generation of anti-tumor immune responses. Here, we present a protocol for characterizing FRCs in human lung cancer. This step-by-step protocol involves enrichment of rare FRCs from tumor tissue for single-cell RNA sequencing and a detailed computational framework for the analysis of FRC-immune cell crosstalk. The topography and physical interactions of FRCs and T cells in specific niche environments are validated using optimized immunohistochemical and high-resolution microscopy approaches. For complete details on the use and execution of this protocol, please refer to Onder et al.1.