Recombinant Alexa Fluor® 488 Anti-Progesterone Receptor antibody [SP2] (ab267523)

ab267523 staining Progesterone Receptor in T47D cells. The cells were fixed with 4% formaldehyde (10 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab267523 at 1/100 dilution (shown in green) and ab190573, Rabbit monoclonal to alphaTubulin (Alexa Fluor^® 647), at 1/250 dilution (shown in ). Nuclear DNA was labelled with DAPI (shown in blue).

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

Overlay histogram showing T47D cells stained with ab267523 (red line). The cells were fixed with 4 % formaldehyde (10 min) and then permeabilized with 0.1 % PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS containing 10 % normal goat serum to block non-specific protein-protein interaction  followed by the antibody (ab267523) (1x 10⁶ in 100µl at 0.1 µg/ml (1/5000)) for 30 min at 22°C.

Isotype control antibody (black line) was Rabbit IgG monoclonal A488 used at the same concentration and conditions as the primary antibody. Unlabeled sample (blue line) was also used as a control.

Acquisition of >5,000 events were collected using a 50 mW Blue laser (488nm) and 530/30 bandpass filter.

IHC image of Progesterone Receptor staining in a section of formalin-fixed paraffin-embedded human breast carcinoma*. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6) in a Biocare Medical NxGen pressure cooker using retrieval settings of 110°C for 20 minutes. Non-specific protein-protein interactions were then blocked in TBS containing 0.025% (v/v) Triton X-100, 0.3M (w/v) glycine and 1% (w/v) BSA for 1h at room temperature. The section was then incubated overnight at +4°C in TBS containing 0.025% (v/v) Triton X-100 and 1% (w/v) BSA with ab267523 at 1/100 dilution (shown in green). Nuclear DNA was labelled with DAPI (shown in blue). The section was then mounted using Fluoromount^®. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). For other IHC staining systems (automated and non-automated), customers should optimize variable parameters such as antigen retrieval conditions, antibody concentrations and incubation times.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre.