single-cell RNA-seq of Merkel cell carcinoma
Cryopreserved, digested MCC tumor samples were thawed and processed following a modified version of the 10x Demonstrated Protocol "Thawing Dissociated Tumor Cells for Single Cell RNA Sequencing" and BioLegend TotalSeq C protocol. The samples were thawed, labeled with TotalSeq C antibodies, and stained with 7-AAD viability staining solution and Apotracker Green (BioLegend). Cells were then subjected to FACS to sort the CD45+ and CD45- cell populations for target enrichment of 2/3 CD45+ cells. After sorting, the final cell preparation was resuspended in RPMI + 10% FBS + SUPERase•In™ RNase Inhibitor (Sigma), filtered, and counted to ensure an optimal concentration of 1e6 cells/mL. Single-cell RNA sequencing libraries were prepared using the 10x Genomics 5' cDNA protocol