Immunohistochemical analysis of parafin-embedded bromodeoxyuridine-labelled chick embryo cells with (MoBu-1) 5-bromodeoxyuridine 

ICC/IF image of ab8039 stained HeLa cells, both BrdU treated (left image) and normal cells (right image). The cells were 100% methanol fixed (5 min) and then subjected to acid hydrolysis using 2M HCL in 0.1% PBS-Tween for 30 minutes at room temperature to denature the DNA. They were  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 (ab8039, 10µg/ml) overnight at +4°C. The secondary antibody (green) was ab96879, DyLight® 488 goat anti-mouse 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. Positive staining can be seen in the BrdU treated cells, but not in the normal cells, demonstrating specificity for BrdU.

IHC image of ab8039 staining, both in normal and BrdU treated rat liver formalin fixed paraffin embedded tissue sections, performed on a Leica Bond

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

Flow cytometry analysis of CEM (human acute lymphoblastic leukemia) cells labelling BrdU with ab8039 at 1 µg/mL. Goat anti-mouse IgG was used as the secondary antibody. The individual cell cycle phases (S, G1, G2/M-phase) are indicated on the figure.