Gene expression of wild-type and Stat3 mutant myeloid cells from EAE mice

Purpose: The goal of this study is to investigate STAT3 signaling in myeloid cells that contribute to antigen-specific T cell activation. This experiment was aimed to determine mRNA transcripts of myeloid cells from preclinical experimental autoimmune encephalomyelitis (EAE) mice and after interacting with autoreactive T cells ex vivo.Methods: Immunomagnetically selected CD11b positive splenocytes from preclinical MOG35-55-EAE mice (9 days post immunization) were co-cultured with CD4+ T cells isolated from naive 2D2 mice in the absence or presence of MOG35-55 peptide (30 microg/ml) for 2 days. CD11b+ myeloid cells in cocultures were then purified by FACS, and single cells captured using the Fluidigm C1 system. Single-cell cDNA samples were prepared on C1 Single-Cell mRNA Seq IFC. Resulting cDNA samples were quantified and normalized for sequencing library preparation utilizing the Illumina Nextera XT Library Preparation Kit. The libraries were normalized and pooled for sequencing on a NextSeq 500 using a 2x75 Mid-output sequencing kit.