Alexa Fluor® 647 Anti-ENO1 抗体 [EPR19758] (ab303584)

AB303584 staining ENO-1 in HeLa (human cervix adenocarcinoma epithelial cell line) 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/0.3 M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with AB303584 at 0.04 µg/ml (shown in red) and ab195883 (Alexa Fluor^®488 Anti-Tubulin antibody [YOL1/34]) at 5 µg/ml (shown in green). Nuclear DNA was labeled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

Immunofluorescence staining of ENO1 in a section of formalin-fixed paraffin-embedded human clear cell carcinoma*.

Performed on a Leica BOND. The section was pre-treated using heat mediated antigen retrieval with EDTA (pH9.0) using retrieval settings of 110°C for 40 minutes. The section was then incubated at room temperature for 1 hour with AB303584 at 1/100 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue). The section was then mounted using Dako Fluorescence Mounting Medium^®.

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.

Flow cytometry overlay histogram showing HeLa cells stained with AB303584 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Triton X-100 for 15 min. The cells were incubated in 1x PBS containing 10% normal goat serum to block non-specific protein-protein interaction followed by the antibody (AB303584) (1x 10⁶ in 100μl at 0.04μg/ml (1/12500)) for 30 min at 22°C.

Isotype control antibody (black line) was Rabbit IgG (monoclonal) Alexa Fluor^® 647 (ab199093) used at the same concentration and conditions as the primary antibody. Unlabeled sample (blue line) was also used as a control.

Acquisition of >5000 events were collected using a 40 mW Red laser (638 nm) and 660/10 bandpass filter.

This antibody gave a positive signal in Hela cells fixed with 4% formaldehyde (10 min) / permeabilized with 0.1% PBS-Triton X-100 for 15 min used under the same conditions.