Single cell RNA and CITE sequencing of cells from synovial tissue biopsies of healthy donors, and Rheumatoid Arthritis (RA) patients with active disease or sustained clinical remission
Single cell RNA and CITE sequencing of cells from synovial tissue biopsies of healthy donors, and Rheumatoid Arthritis (RA) patients with active disease or sustained clinical remission
Synovial biopsies were taken using 14G Precisa Needle (HS Hospital Service, Italy) in ultrasound-guided protocol, and digested as described previously (Alivernini et al, 2020). Live CD45+ cells from the synovial tissue (ST) of healthy donors (n=3), patients with active RA (n=5) and patients in sustained remission (n=2) were sorted for single cell RNA (scRNA) or CITE (scCITE) sequencing using the 10x Genomics platform. Maximum 20,000 immune cells from tissue were sorted into Protein LoBind 1.5 ml Eppendorf tube containing 300 μl of RPMI media with 10% of FSC. Cells were loaded onto a Chromium Controller (10X Genomics) for single-cell partitioning, followed by library preparation using Single-Cell 3' Reagent Kits v3.1. For 3 healthy ST and 4 ST from active RA (Figure S1), Totalseq Hashtag were used (Biolegend #394601, #394603, #394605, #394607, #394609) to combine 2 samples per run, and TotalSeq™-A Human Universal Cocktail (V1.0) was used to collect protein expression (CITEseq) data together with transcriptome data.  As recommended by 10X Genomics protocol for the 10X Single Cell 3' Reagent Kit v3.1 As recommended by 10X Genomics protocol for the 10X Single Cell 3' Reagent Kit v3.1