Anti pkm

Porcine kidney-15 (PK-15) and human embryonic kidney (HEK293T) cells were cultured at 37°C and 5% CO₂ in Dulbecco’s modified Eagle’s medium (DMEM; Gibco) supplemented with 10% fetal bovine serum (FBS; Gibco), 100 IU/mL penicillin, 100 g/mL streptomycin, and 2 mM L-glutamine (Gibco). The SVA strain, CH-GDFS-2018, was isolated by our group and propagated and titrated in PK-15 cells using the Reed–Muench method [46 (link)]. The SVA strain was incubated in a water bath at 70°C for 2 h to generate a heat-inactivated SVA (Heat-SVA).
Sodium oxamate, 2-deoxyglucose (2DG), sodium lactate, dichloroacetate (DCA), and glucose were purchased from Macklin (Shanghai, China). The 2DG (50 mM), sodium oxamate (25 mM), DCA (5 mM), sodium lactate, and glucose were directly solubilized in DMEM or phosphate buffered saline (PBS) at the indicated concentrations. DMEM, PBS, and FBS were obtained from Gibco (Grand Island, NY, USA). Anti-HIF-1α, Anti-PGK1, anti-PKM, anti-GAPDH, anti-RIG-I and anti-β-actin primary antibodies were purchased from Santa Cruz (Shanghai, China). A primary antibody of rabbit anti-VP2 of SVA was prepared and stored in our laboratory [27 (link)]. The respective horseradish peroxidase (HRP)-conjugated secondary antibodies were obtained from Abcam (Cambridge, UK).

Cell lysates were prepared and western blots were performed using the same procedures as previously described [18 (link)]. Antibodies used were anti-PKM (Santa-Cruz, clone C-11, 1 : 1000), anti-PGP (Santa-Cruz, clone E10, 1 : 1000), anti-β-actin (Sigma, clone AC-15, 1 : 5000), anti-p53 (clone DO1, Santa-Cruz, 1 : 1000), anti-phospho-p53 (Cell Signalling, 1 : 1000), anti-P-H2AX (Millipore, 1 : 1000) and anti-tubulin (Sigma, 1 : 40 000).