GSM7990493: C92_scRNAseq_11_Upper_Carious_Pulp; Homo sapiens; RNA-Seq
Teeth were collected immediately after extraction and placed in sterile 0.9% NaCl, on ice. Dental pulp was obtained from carious teeth following the protocol we previously applied for the isolation of healthy human pulp tissues.66 Briefly, each tooth was cracked with a press, and carefully opened with forceps. Isolated dental pulp was placed in a Petri dish filled with cold HBSS and minced into smaller pieces (<2 mm diameter) for easier dissociation. The dental pulp pieces were transferred to falcon tubes and dissociated using 5 U/mL Collagenase P (11 213 873 001, Sigma-Aldrich, Buchs, Switzerland) in PBS for 40 min at 37 °C. After incubation in the enzymatic solution, tissue pieces were further mechanically disaggregated by pipetting and centrifuged at 300 × g at 4 °C for 10 min. Cell pellet was resuspended in HBSS containing 0.002% Bovine Serum Albumin (BSA; 0163.2, Roth AG, Arlesheim, Switzerland) and filtered through a 70 μm cell strainer. Cells were loaded into the 10X Chromium Next GEM Chip G, aiming the recovery of ~10000 cells per sample. scRNAseq libraries were generated according to 10X Chromium Next Gem Single Cell 3' Reagent Kits v3.1 (CG000204 Rev D) and sequenced on the Illumina Novaseq 6000 system