GSM9405282: Foam Cell 1; Homo sapiens; RNA-Seq
THP-1 cells (#TIB-202, ATCC;) were cultivated in RPMI-1640 medium (#10-041-CV, Corning;) with 10% FBS. They were then treated with PMA (#5.00582.0001, Millipore Sigma) to reach 15 nM concentration and plated in a 12-well plate (#150628, Thermo Fisher Scientific) at a density of 5 x 105 cells per well. The plate was incubated for a duration of 3 days to promote differentiation. On the fourth day, the cells were gently washed with 1 mL of fresh medium to remove any residual PMA and other soluble factors. Following this, oxLDL (#L34357, Invitrogen) was introduced to each well to reach a final concentration of 50 µg mL-1, ensuring thorough mixing for uniform exposure. The cells were incubated for an additional four days to allow for foam cell formation. Upon completion of the incubation period, the medium was carefully aspirated from each well. The cells were detached by adding 200 µL of trypsin-EDTA (#25-053-CI, Corning) per well and incubating at 37°C for 5 min to facilitate cell detachment. To neutralize the trypsin activity, 1 mL of medium was added per well and the mixture was gently pipetted to ensure complete suspension of the cells. Finally, the cell solution was filtered through a 100 µm nylon cell strainer (#352360, Falcon) to remove any cell clumps, thereby obtaining a uniform single-cell suspension ready for further analysis. Library preparation was conducted with Chromium GEM-X Single Cell 3' Reagent Kits v4 (10x Genomics). Libraries were sequenced on NovaSeq X Plus.