Rabbit anti actin primary antibody

Cal33 cells were treated with DMSO, 500 nM or 1 μM albendazole for 24 hours. As a positive control for apoptosis, Cal33 cells were treated with 1μM staurosporin (Selleck Chemicals) for 4 hours. PARP primary antibody (Genetex Inc., cat. GTX100573), was administered according to manufacturer recommendations. Actin levels were detected using a rabbit anti-actin primary antibody (Sigma-Aldrich, cat. A2066), and the same secondary antibody (G-anti-R) and detection method as previously noted. Western blot image is presented as scanned.

We used the GAL4-UAS system for RNAi and the fly lines include UAS-Dcr2-Nubbin-GAL4 (a gift from Dr Osamu Shimmi, Institute of Biotechnology, University of Helsinki, Finland) and the following UAS lines: FBst0026777 (Chinmo-1) and FBst0033638 (Chinmo-2) for Chinmo, FBst0032347 for exportin 6. These are also described in flybase (http://flybase.org). W1118 was a kind gift from Minna Poukkula (Institute of Biotechnology, University of Helsinki, Finland). Wing discs were dissected in PBS and fixed with 4% paraformaldehyde for 30 min at room temperature. Tissues were permeabilized in 0.5% Triton X-100 in PBS (PBT) for 10 min and blocked with 5% bovine serum albumin (BSA) in PBT for 1 h. Tissues were incubated with rabbit anti-actin primary antibody (Sigma) (A2103, dilution 1:250) overnight at 4°C. Tissues were washed in PBT for 1 h, and then incubated with Alexa-Fluor-labeled rabbit secondary antibodies and DAPI for 1 h at room temperature. Tissues were washed with PBT for 1 h and mounted in glycerol containing n-propylgallate (2.5%). Images were acquired using the Leica TCS SP5 confocal microscope, with a 63×1.3 NA objective and LAS AF software. Nuclear and cytoplasmic fluorescence intensities were measured using CellProfiler.