Primary antibodies for caspase 3

Analysis of apoptotic HL-60 cells was performed with the CBMN assay. Cell cultures were established as in micronucleation study. Cells were treated with DOX at two different concentrations, 0.5 and 2.0 μg ml⁻¹ for 6 h. Slides were stained with May-Grünwald and 10% Giemsa. Standard criteria were used for the identification of apoptotic cells [14 ].
Complementary to the above, caspase-3-level-Western blotting analysis was achieved. HL-60 cells were treated with DOX at the same concentrations and time as above. The procedure was followed according to recent publication [24 (link)]. Caspase-3 and β-actin were detected with primary antibodies for caspase-3 (1:1000, Cell Signaling, Boston, MA, USA, 9665) and β-actin (1:1000, Cell Signaling, Boston, MA, USA, 4967). The protein amount in the bands of each lane was quantified by densitometric analysis relative to the actin numerical quantities. In each independent experiment, protein amount variation fold was calculated considering equal to 1 the protein/β-actin ratio observed in untreated cells [25 (link), 26 (link)].

Primary antibodies for α-SMA, Rb, SKP2, P27 were purchased from Santa Cruz Biotechnology (USA). Primary antibodies for Caspase3, P-Rb, P-ERK were purchased from Cell Signaling Technology (USA). All of the secondary antibodies were purchased from Santa Cruz Biotechnology (USA). Recombinant human TGF-β1 and staurosporine (STS), a positive apoptosis stimulus, were obtained from Sigma (USA).