Streptomycin penicillin neomycin solution

PANC1 human pancreatic ductal adenocarcinoma cell line (obtained from the American Type Culture Collection) was cultured in Dulbecco’s modified eagle’s medium (DMEM, Sigma-Aldrich, Israel) supplemented with 10% fetal bovine serum (FBS, Biological Industries, Israel), 1% L-glutamine, 1% sodium pyruvate, and 1% streptomycin-penicillin-neomycin solution (Biological Industries). The cells were used within 6 months of resuscitation, and were periodically tested to be mycoplasma-free. Cells were cultured at 37 °C in a humidified atmosphere containing 5% CO₂.

Murine pancreatic adenocarcinoma cell line Panc02 was kindly provided by Dr. Q Yao (Baylor College of Medicine, Houston, TX, USA). Murine colon carcinoma cell line MC38 was a kind gift from Dr. John C. Morris (National Cancer Institute, Bethesda, MD, USA). These cell lines were maintained in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% (v/v) FBS, 1% L-glutamine, 1% sodium pyruvate, and 1% streptomycin-penicillin-neomycin solution (Biological Industries, Cromwell, CT, USA). A recombinant lentiviral PD-L1 vector that coexpresses PD-L1 and YFP (yellow fluorescent protein) was used to transduce these cells to establish PD-L1-expressing stable tumor cell lines, Panc02-PD-L1 and MC38-PD-L1. These tumor cells, which stably express human PD-L1, were maintained in the media supplemented with 500 ng/mL or 200 ng/mL puromycin (Invitrogen, Carlsbad, CA, USA).

PyMT murine breast carcinoma cell line was obtained as described in Chang et al.17 (link) EMT6 murine breast carcinoma and HEK-293T human embryonic kidney cell lines were purchased from ATCC (Manassas, Virginia, USA). Mouse embryonic fibroblast cell line was obtained as described in Weidenfeld-Baranboim et al.18 (link) Cell lines were cultured in Dulbecco’s modified eagle’s medium (Sigma-Aldrich, Israel) supplemented with 10% fetal bovine serum 1% L-glutamine, 1% sodium pyruvate, and 1% streptomycin-penicillin-neomycin solution (Biological Industries, Israel). All primary cells (lymphocytes, MDSCs, bone marrow cells, macrophages) were cultured in Iscove’s modified Dulbecco’s medium supplemented as described above. Cells were routinely tested to be mycoplasma-free.