GSE158579
CNS-Native Myeloid Cells Drive Immune Suppression in the Brain Metastatic Niche through Cxcl10 [R10_PD-L1+VISTA treatment]
We performed CITE-seq (10x Genomics-based) to profile and compare the transcriptomes and cell surface expression of immune epitopes in the brains of anti-VISTA+anti-PD-L1 treated (Dual TX) and control, non-treated mice. We sequenced a total of eight  samples (4 control, 4 dual TX). 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: Control/Non-treated (2) HTO2: Control/Non-treated ; (3) HTO3: Control/Non-treated ; (4) Control/Non-treated ; (5) HTO5: Dual TX; (6) Dual TX; (7) HTO7: Dual TX; (8) HTO8: Dual TX. The following sample comparisons were made: HTO1 - HTO4 were compared to HTO5 - HTO8. Overall design: Metastasis-burdened brains were obtained from 2-3 month old female mice with or without anti-VISTA + anti-PD-L1 treatment (Dual TX). These mice were administered Dual TX every other day starting 3 days-post metastasis injection. Brain metastasis was established by carotid injection of E0771 cells. Cell suspensions were prepared by percoll gradient centrifugation enrichment to obtain suspensions enriched for immune cells from the brain. 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.