Fitc conjugated mouse anti rabbit igg antibody

After treated for 48 h, 2 x 10⁵ NALM-6/R cells were subjected to apoptosis assay using an Annexin V Apoptosis Detection Kit-APC (eBioscience Company, USA), following the manufacturer's instructions. For the mitochondrial membrane potential assessment, after different treatments for 48 h, 2 x 10⁵ NALM-6/R cells were analyzed using a JC-1 fluorescent probe kit (Beyotime Company, China), following the manufacturer's instructions. To measure the intracellular reactive oxygen species (ROS) levels, about 2 x 10⁵ NALM/R cells subjected to different treatments were washed in PBS buffer twice, and then incubated in 1 ml of serum-free RPMI 1640 medium containing 10 μM of H₂DCFDA for 30 min at 37°C. The cells were harvested, and then washed in serum-free RPMI 1640 medium buffer twice to remove the remaining H₂DCFDA. The fluorescent intensity was measured by flow cytometry. For assessing the degree of DNA damage, 2 x 10⁵ NALM-6/R cells were incubated for 15 min on ice in hybridization buffer (PBS containing 0.5% bovine serum albumin (BSA) and 0.25% Triton X-100). After centrifugation, the cells were incubated with rabbit monoclonal anti-γH2A.X antibody (Cell Signaling Technology, USA) for 1 h, then washed with PBS and incubated with an FITC-conjugated mouse anti-rabbit IgG antibody (BD Pharmingen) for 30 min in the dark at room temperature.

CD34⁺CD38⁻ KG1α cells were exposed to 20 or 40 nM IDA in combination with or without 0.75 μM chidamide for 24 h, with an untreated group as the control. Cells were harvested and incubated for 15 min on ice in a hybridization buffer (PBS containing 0.5% bovine serum albumin (BSA) and 0.25% Triton X-100). After centrifugation, the cells were incubated with rabbit monoclonal anti-γH2A.X antibody (Cell Signaling Technology, USA) for 1 h, then washed with PBS and incubated with an FITC-conjugated mouse anti-rabbit IgG antibody (BD Pharmingen) for 30 min in the dark at room temperature. The stained cells were analyzed by flow cytometry (FACS C6, BD Biosciences).