title: GSM4760976: TAMs - 8 weeks - an1; Mus musculus; RNA-Seq
library_construction_protocol: Mammary glands were collected in DMEM on ice then minced. Tissue was digested in two 30 min rounds of collagenase and DNase with agitation at 37C. At the end of each round, digested tissue was washed through a 100um cell strainer on ice. Red blood cells were lysed with ACK buffer and remaining cells washed and counted. Cells were stained with anti-mouse CD45, anti-mouse CD11b, anti-mouse Ly6C, anti-mouse CD14, anti-mouse I-A/I-E, anti-mouse CD11c, anti-mouse CD24, anti-mouse CD29, anti-mouse CD31, and Zombie NIR live/dead. Sorted macrophages were CD45+, MHCII+, CD14+. Detailed antibody information and gating strategy included in manuscript. Sorted cells were pelleted and snap frozen on dry ice. RNA was harvested using Trizol and measured on a spectrophotometer and run out on a Bioanalyzer to confirm quality. Libraries were prepared according to manufacturer instructions using Nugen Ovation plus Nextera (<100 ng RNA)