title: scRNA-seq of human intestinal TRMs as well as intestinal organoids with autologous tissue-resident or blood derived immune compartment from 3 distinct donor samples
design: scRNA-seq of human intestinal TRMs as well as intestinal organoids with autologous tissue-resident or blood derived immune compartment from 3 distinct donor samples
library_construction_protocol: All the samples were dissociated as follows: matrigel domes containing organoids in coculture with immune cells or immune cells alone were dislodged and mechanically broken up and transferred to 1% BSA-coated tubes. The dome fragments were centrifuged at 400g, 4min, RT. The supernatant was removed and the enzymes of the neural tissue dissociation kit (Miltenyi Biotec) were mixed in HBSS-1%BSA buffer. The matrigel containing the cells were dissociated to single cells for a total of 30 min with thorough pipetting every 7 minutes. Next, cells were filtered through a 40 µM filter and single cells were centrifuged at 450g, 4min, RT. Dead cells were magnetically removed and the viable cell pellet was subsequently resuspended in DPBS 1% BSA. Nuclei acid extraction protocol was performed according to the Chromium Next GEM Single Cell 3' Reagent Kits v3.1 (Dual Index), CG000315, Rev E.  The kit used was: Chromium Next GEM Single Cell 3' Kit v3.1, 16 rxns PN-1000268. Library Construction protocol was performed according to the Chromium Next GEM Single Cell 3' Reagent Kits v3.1 (Dual Index), CG000315, Rev E.  The kit used was: Library Construction Kit 16 rxns PN-1000190.