GSM8819513: SP4; Mus musculus; RNA-Seq
Fresh liver tissue samples were cut into 1 mm3 pieces and digested in HBSS (Gibco, USA) buffer containing 0.05% collagenase IV (Invitrogen, Carlsbad, USA) and 0.002% DNase I (Invitrogen, Carlsbad, USA) at 37°C with gentle agitation for 30 minutes. The dissociated cells were filtered through 70 μm sterile cell strainers. Immune cells were then separated by density gradient centrifugation with 36% Percoll medium (Sigma-Aldrich, St. Louis, USA), and residual erythrocytes were removed using a red blood cell lysate solution (Sigma, USA). The immune cells were subsequently re-suspended in phosphate-buffered saline (PBS) to create a single-cell suspension. The single-cell suspension was loaded onto the MobiNova®-100 chip with 3' transcriptome microspheres, droplet-generating oil, and reverse transcription reagents. RNA transcripts from individual cells were uniquely barcoded and reverse-transcribed within the droplets formed on the chip. Following this, pooled cDNA was purified and enriched through PCR amplification to construct cDNA libraries.