Chemotherapy-mediated changes in the immune cell landscape of murine triple negative breast cancer.

We performed CITE-seq (10x Genomics-based) to profile and compare the transcriptomes and cell surface expression of immune epitopes in the primary breast tumors of transgenic mice (C3-1-TAg) with or without MCT chemotherapy. We sequenced a total of six different tumor samples. We created an antibody pool consisting of 35 different antibodies and stained each sample individually with this antibody pool. Then, we stained each sample with it's own unique hashing antibody so that we could subsequently pool the samples for loading onto the 10x Chromium and later prepare one library consisting of all sight samples and finally separate each sample in silico by it's unique hashing antibody. The samples are as follows: (1) HTO1: tumor of PBS treated C3-1-TAg mouse; (2) HTO2: tumor of PBS treated C3-1-TAg mouse; (3) HTO3: tumor of PBS treated C3-1-TAg mouse; (4) HTO4: tumor of MCT treated C3-1-TAg mouse; (5) HTO5: tumor of MCT treated C3-1-TAg mouse; (6) HTO6: tumor of MCT treated C3-1-TAg mouse. The following sample comparisons were made: HTO1, HTO2, and HTO3 versus HTO4, HTO5, and HTO6. Breast tumors were isolated from C3-1-TAg mice treated with either MCT or PBS for 4 weeks. These mice were started on treatment when the tumors were around 5 mm* 5 mm in size. Tumors were dissociated using collagenase enzyme and the single cells were enriched with CD45+ immune cells using MACS magnetic beads (Miltenyi). We created an antibody pool consisting of 35 different antibodies and stained each sample individually with this antibody pool. Then, we stained each sample with it's own unique hashing antibody so that we could subsequently pool the samples for loading onto the 10x Chromium and later prepare one library consisting of all eight samples and finally separate each sample in silico by it's unique hashing antibody.