Stable Expression of CXCR3 Isoforms in HEK293 Cells

HEK293 cells were grown at 37°C in Dulbecco’s Modified Eagle (DMEM) media (Corning Inc., Corning, NY) supplemented with 10% fetal bovine serum (FBS, Corning Inc.) and 1% penicillin/streptomycin/amphotericin B (Corning Inc.). cDNA constructs containing CXCR3A (GenBank accession # NM_001504; GenScript) and CXCR3B (GenBank accession # NM_001142797; GenScript) were purchased from GenScript. Prior to transfection, HEK293 cells were seeded at the appropriate density to ensure 75% cell confluence within 24 hrs. Transfection of the cDNA constructs was performed after 24 hrs using X-tremeGENE Transfection Reagent (Roche Holding AG, Basel, Switzerland) according to the manufacturer’s protocol using a 3:1 transfection reagent to cDNA ratio as previously described.[35 (link)] Following 48 hrs of transfection, the media was changed to complete media containing the selection agent G418 (400 μg/mL, Corning Inc.). The cells were maintained in G418 containing media to generate stable cell lines, which were used in subsequent experiments.