Histone h3k36me3 antibody

Protein isolation and Western blots were performed using standard protocols as described elsewhere^{41 (link)}. The following primary antibodies were used: R2 (Santa Cruz Biotechnology, Dallas, USA, cat. #: sc-10844, 1:1000), Wee1 (Santa Cruz Biotechnology, cat. #: sc-9037, 1:1000), Anti-Histone H3.3 G34W (RevMAb Biosciences, South San Francisco, USA, cat. #: 31-1145-00, 1:1000), Histone H3K36me3 antibody (Active Motif, Cambridge, UK, cat. #: 61021, 1:1000), Cdc2 p34 (Santa Cruz Biotechnology, cat. #: sc-54, 1:500), Phospho-cdc2 Tyrosine 15 (Cell Signaling, Denver, USA, cat. #: #9111, 1:500), and Cyclin B1 (Santa Cruz Biotechnology, cat. #: sc-245, 1:200). As secondary antibody we used: Anti-Rabbit IgG (Sigma-Aldrich, cat. #: A-9169,1:10000), rabbit anti-goat IgG-HRP (Santa Cruz Biotechnology, cat. #: sc-2768, 1:10000) and goat anti-mouse IgG-HRP (Santa Cruz, cat. #: sc-2005, 1:5000). Anti-β-Actin antibody (Sigma-Aldrich, cat. #: A2228, 1:100000) or Anti-β-Tubulin (Sigma-Aldrich, cat. #: T4026,1:2000) were used to detect β-actin or β-tubulin as a housekeeping protein.

The alkaline phosphatase/RED detection system (Dako, cat. #: K5005) was used for immunohistochemistry and immunochemistry on formalin-fixed and paraffin-embedded tissue or cell pellets via the avidin-biotin-complex-method. The following antibodies were used: Anti-Histone H3.3 G34W (RevMAb Biosciences, cat. #: 31-1145-00, 1:400), Wee1 (Santa Cruz Biotechnology, cat. #: sc-037, 1:25), Cdc2 p34 (Santa Cruz Biotechnology, cat. #: sc-54, 1:200), Cyclin B1 (Santa Cruz Biotechnology, cat. #: sc-245, 1:25), Histone H3K36me3 Antibody (Active Motif, cat. #: 61021, 1:8000), R2 (Santa Cruz Biotechnology, cat. #: sc-10844, 1:100), and Ki-67 Antigen (Dako, cat. #: M7240, 1:200). As retrieval methodes all sides were pretreated by a steamer in EDTA pH 8. Consensus evaluation was done on multihead microscope (CL, TFB, PM).