LM22 is a signature matrix file consisting of 547 genes that accurately distinguish 22 mature human hematopoietic populations isolated from peripheral blood or in vitro culture conditions, including seven T cell types, naïve and memory B cells, plasma cells, NK cells, and myeloid subsets. LM22 was designed and extensively validated on gene expression microarray data, but is also applicable to RNA-Seq data for hypothesis generation (section 5.1). Here, we illustrate how to prepare Affymetrix microarray data for use with LM22, and how to run CIBERSORT with LM22 to characterize the leukocyte composition of prostate biopsies obtained from patients with prostate cancer and from healthy subjects. To follow the examples in this section, download GSE55945 CEL files from GEO (https://www.ncbi.nlm.nih.gov/geo/download/?acc=GSE55945&format=file). Processed data for GSE55945 can be downloaded from the CIBERSORT website.