T cell activation experiments require fresh, high-purity CD3⁺ T cells with the cell density adjusted to approximately 1×10⁶ cells/mL. CD3/CD28 dual-signal stimulation is used for activation with appropriate IL-2 supplementation. Strict aseptic operation should be implemented throughout the process to prevent mycoplasma contamination, and cell culture is maintained at 37 ℃ with 5% CO₂. The detection time point needs to be precisely controlled to avoid apoptosis induced by excessive activation; meanwhile, blank controls and isotype controls shall be set up.