GSM6858205: IgE-CAI skin lesion (hashtag library); Mus musculus; RNA-Seq
On days 3 and 5 post-challenge, excised ears were cut into small pieces by using razors, and were incubated with Liberase TM (0.25 mg/mL; Roche) in RPMI complete medium at 37 °C for 50 min. The cells were then subjected to density separation with 25% and 65% Percoll PLUS (Cytiva) and cells were recovered from the interphase, to remove dead cells and red blood cells. Recovered cells were subsequently incubated with TotalSeq anti-mouse Hashtag antibodies (clone M1/42, 30-F11; BioLegend). 10,000 labeled cells were trapped and reverse-transcribed using BD Rhapsody (BD) according to the manufacturer's instructions, followed by the preparation of cDNA libraries and hashtag libraries by TAS-Seq method (Shichino et al. Commun Biol 2022). The sequencing analysis were performed  a Novaseq 6000 sequencer and a Novaseq S4 200 cycles v1.5 kit (Illumina).