Cancer-infecting virus 'warms up' cold tumors and improves immunotherapy JExpMed
) was used. The pSC65 cloning plasmid was remade using Gibson cloning so that firefly luciferase was expressed from the pSE/L promoter and the dnTGFβmm with an IL-2 signal sequence was expressed from the p7.5 early/late promoter. This was recombined into the TK gene using Gibson cloning .. In brief, this is mini-monomer binds TGFβRII with the same affinity as TGFβ1 and TGFβ3 dimers; however, it cannot recruit TGFβRI.at an MOI of 0.1 for 2 h in DMEM supplemented with 10% FBS .
Cell surface staining was performed on ice in PBS for 20 min with surface antibodies and Zombie Viability Dye . Cells were then washed in PBS and either run for live panels or fixed with 4% paraformaldehyde for 5 min at room temperature and then washed in PBS. For phospho staining, cells were fixed with ice-cold 90% methanol at −20°C for 20 min, then washed with permeabilization wash buffer , and stained overnight at 4°C.
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