Anti-Fascin antibody [EP5902] (ab126772)

All lanes : Anti-HLA-C antibody [EPR6749] (ab126722) at 1/1000 dilution

Lane 1 : Wild-type HAP1 whole cell lysate
Lane 2 : FSCN1 (Fascin) knockout HAP1 whole cell lysate
Lane 3 : HepG2 whole cell lysate

Lysates/proteins at 20 µg per lane.

Predicted band size: 54 kDa

Lanes 1 - 3: Merged signal (red and green). Green - ab126772 observed at 54 kDa. Red - loading control, ab9484, observed at 37 kDa.

ab126772 was shown to specifically react with Fascin in wild-type HAP1 cells as signal was lost in FSCN1 (Fascin) knockout cells. Wild-type and FSCN1 (Fascin) knockout samples were subjected to SDS-PAGE. Ab126772 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

All lanes : Anti-Fascin antibody [EP5902] (ab126772) at 1/10000 dilution

Lane 1 : SH SY5Y cell lysate
Lane 2 : K562 cell lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : HRP-conjugated goat anti-rabbit at 1/2000 dilution

Predicted band size: 54 kDa

Immunocytochemistry/Immunofluorescence analysis of U87-MG (human glioblastoma) labelling Fascin with purified ab126772 at 1/500. Cells were fixed with 100% methanol. An Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (ab150077) at dilution of 1/1000 was used as the secondary antibody. Nuclei couterstained with DAPI (blue). 

Secondary Only Control: PBS was used instead of the primary antibody as the negative control. 

ab126772, at 1/250 dilution, staining Fascin in paraffin-embedded Human Hodgkin's lymphoma tissue by Immunohistochemistry.

Overlay histogram showing HeLa cells stained with ab126772 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab126772, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1μg/1x10⁶ cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.

ab126772, at 1/250 dilution, staining Fascin in paraffin-embedded Human colon tissue by Immunohistochemistry.

ab126772 showing positive staining in Normal tonsil tissue.

ab126772 showing positive staining in Glioma tissue.

ab126772 showing positive staining in Normal kidney tissue.