Pe anti ki67 antibody

7.5 × 10⁴ Huh7 or HT-29 cells were seeded in 1 mL of complete growth medium in 24-well and cultured for 24 h until they had fully adhered. PBMCs from HCC or CRLM patients or HD were stimulated 16 h with IL-2 (50 ng/mL) (Miltenyi Biotec) in complete medium. Subsequently, PBMCs (7.5 × 10⁵ cells/well) were added to tumor cells in a final volume of 2 mL (10:1 ratio) and incubated at 37 °C and 5% CO2 for 48 h. After 48 h, PBMCs were eliminated and adherent Huh7 or HT-29 cells were detached. Cell viability was analyzed by 7-AAD staining (Miltenyi Biotec). After fixation and permeabilization with commercially available kit (Transcription Factor Buffer Set, BD Pharmingen), tumor cells were stained with PE-anti-Ki67 antibody (BD Bioscience, San Jose, CA, USA). The samples were analyzed by flow cytometry.