title: GSM8209879: MedullaryTumor_enzymatic_1; Homo sapiens; RNA-Seq
library_construction_protocol: Single cells were isolated from tissue samples using and enzymatic dissociation method. Approximately 5-250 mg of fresh tissue samples were washed in HBSS, thoroughly minced on ice, and placed in dissociating solution at 37°C with gentle pipetting every 5 minutes.  Tissue samples were dissociated with 25-30 ul enzyme D Multi Tissue Dissociation Kit (MTDK) (Miltenyi Biotec) or enzyme A Neural Tissue Dissociation Kit P (NTDK) (Miltenyi Biotec) or 2 mg/ml collagenase IV (Gibco, Thermo Fisher Scientific) or collagenase I (Gibco, Thermo Fisher Scientific) in 870 mM HBSS, 10% FBS, 20 mM HEPES for 20-30 minutes. Obtained homogenate was filtered in 3-5ml of Wash Buffer (1x DPBS, containing 10% FBS, 20mM HEPES, 6mM glucose) through a pre-wetted 70μm cell culture filter (Miltenyi Biotec) and centrifuged for 5 minutes at 300xg (4 °C).  After all, the pellet was resuspended in a Wash Buffer volume of 100-400 ul, depending on the pellet size. Single cells were captured and barcoded, and cDNA libraries were generated using the Chromium Next GEM Single Cell 3ʹGEM, Library & Gel Bead Kit v3.1 (10X Genomics) according to the manufacturer's instructions. Final libraries were multiplexed and sequenced on an Illumina Novaseq 6000 / HiSeq2500 platform, using the S4 Reagent Kit v1.5.