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The total protein isolated from Jurkat cells were separated using radioimmunoprecipitation assay (RIPA) buffer (Beyotime Biotechnology, Shanghai, China). The protein concentration was determined using a bicinchoninic acid protein assay kit (Wanleibio, Shenyang, China). Furthermore, proteins were separated using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE; Wanleibio, Shenyang, China) and electrophoretically transferred to polyvinylidene fluoride (PVDF) membranes (Millipore, Billerica, MA, USA). Subsequently, they were blocked with 5% nonfat milk for 1 h at room temperature and incubated overnight at 4 °C with IL-7 (1:1000; ABclonal, Wuhan, China) and GAPDH primary antibodies (1:1000; Wanleibio, Beijing, China). The membranes were washed with Tris-buffered saline (TBS) and Tween-20 (TBST) and incubated with goat anti-rabbit secondary antibodies for 2 h at room temperature. Subsequently, the protein blots were visualized using an enhanced chemiluminescence reagent (ECL; Wanleibio, Beijing, China), while GAPDH was used as the internal reference. The intensity was quantified using the ImageJ software (National Institutes of Health, Bethesda, MD, USA).

The cells were harvested in RIPA Lysis Buffer and lysed using ultrasound (Wanleibio, Shenyang, China). BCA Reagent was used to determine total protein content (Wanleibio, Shenyang, China). SDS-PAGE (Wanleibio, Shenyang, China) was used to separate equivalent quantities of protein extract, which was then deposited onto PVDF membranes (Millipore, USA). Cleaved-Caspase-1 (Wanleibio, Shenyang, China), cleaved-Caspase-3 (Wanleibio, Shenyang, China), cleaved-Caspase-4 (Affinity Biosciences, Suzhou, China), cleaved-Caspase-8 (Affinity Biosciences, Suzhou, China), GSDMD (Affinity Biosciences, Suzhou, China), GSDME (ABclonal, Wuhan, China), and GSDMD-N (Affinity Biosciences, Suzhou, China) were the primary antibodies employed in this test. After blocking with 5% skim milk for an hour, the membranes were incubated overnight with primary antibodies at 4°C. The membranes were then incubated with HRP-conjugated secondary antibodies (Wanleibio, Shenyang, China) and detected using an enhanced chemiluminescence substrate kit (Wanleibio, Shenyang, China) after washing.