Recombinant Anti-Topoisomerase II alpha + Topoisomerase II beta/TOP2B antibody [EPR5377] (ab109524)

Blocking and diluting buffer and concentration: 5% NFDM/TBST.

TOP II alpha recombinant protein (aa 1-220) containing a his-tag.

ab109524 staining Topoisomerase II alpha + Topoisomerase II beta in the human cell line HeLa (human cervix adenocarcinoma) by intracellular flow cytometry. Cells were fixed with 4% paraformaldehyde, permiabilised with 90% methanol and the sample was incubated with the primary antibody at a dilution of 1/20. A goat anti rabbit IgG (Alexa Fluorr^® 488) at a dilution of 1/2000 was used as the secondary antibody. 

Isoytype control: Rabbit monoclonal IgG (Black) 

Unlabelled control: Cell without incubation with primary antibody and secondary antibody (Blue) 

ICC/IF image of ab109524 stained HeLa cells. The cells were 4% Formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab109524, 10µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

ab109524, at a 1/100 dilution, staining Human Topoisomerase II alpha + Topoisomerase II beta in breast tissue, using Immunohistochemistry, Formalin/PFA-fixed paraffin-embedded tissue.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.