title: GSM9242499: immune cells, PFOA treated, uninfected; Mus musculus; RNA-Seq
library_construction_protocol: Half of the spleen tissues were mechanically dissociated and passed through 70 µM cell strainer for preparing single cell suspensions. Red blood cells were lysed using Red Cell Lysis Buffer (Sigma-Aldrich, St. Louis, MO) for 5 minutes at room temperature. After washing, the cells were resuspended in complete RPMI 1640 medium containing 10% FBS 2 × 10⁶ cells were incubated overnight for probe hybridization in a Bio-Rad T100 thermocycler. For single-cell encapsulation, approximately 10,000 cells were loaded onto the 10x Chromium system (10X Genomics), where Gel-Bead-in-Emulsions (GEMs) were generated. The GEM incubation reaction was carried out in the Bio-Rad T100 thermocycler to facilitate the encapsulation of individual cells with their corresponding beads for subsequent RNA capture. Probe-labeled cDNA was then recovered from GEMs by pre-amplification PCR, followed by DNA cleanup to remove any impurities. The amplified cDNA underwent 10x Genomics-specific sample indexing according to the manufacturer's protocol, followed by post-indexing cleanup with the SPRIselect reagent. Library quality was assessed by quantifying and evaluating the size distribution of the libraries using an Agilent Bioanalyzer. Libraries were then pooled in equimolar amounts for sequencing on a NovaSeq instrument (Illumina).
identifiers.namespace: GEO
identifiers.id: GSM9242499_r1