GSM5550104: Psoriasis04_postTx_week12_LS; Homo sapiens; RNA-Seq
6 mm punch skin biopsy tissues from human skin were bisected, and one of the two bisected skin tissues was incubated in culture medium for harvesting emigrating cells. Harvested skin tissue was immediately placed in 0.2% Dispase II (Sigma-Aldrich) and incubated in a humidified incubator at 37 Celsius and 5% CO2 for 3 hours. Then, epidermis and dermis were separated with forceps and sliced into small pieces with #10 blade scalpels. Epidermis and dermis were separately cultured in RPMI-1640 medium with L-glutamine (Cytiva) supplemented with 10% human albumin serum (Sigma-Aldrich) and without any enzyme or antibiotic agent in a humidified incubator at 37 Celsius and 5% CO2. Nonplastic adherent cells that had emigrated out of epidermis and dermis were harvested after 48 hours. The harvested cells from epidermis and dermis were filtered through a 40-µm cell strainer (Corning) and stored on ice. The 10x Genomics Chromium Single Cell 3′ Reagents Kit user guide (https://support.10xgenomics.com/single-cell-gene-expression/library-prep) was used to prepare the single cell suspension. The appropriate volume of each sample was diluted to recover 10,000 cells. Subsequently, the single cell suspension, Gel Beads and oils were added to the 10x Genomics single-cell chip. After droplet generation, samples were transferred into PCR tubes and we performed reverse transcription using a ProFlex PCR Thermocycler System (Applied Biosystems). After reverse transcription, cDNA was recovered using a recovery agent, provided by 10x Genomics, followed by silane DynaBead clean-up as outlined in the user guide.