Immunocytochemistry/ Immunofluorescence analysis of Jurkat (Human T cell leukemia T lymphocyte) cells labeling Staufen/STAU1 with Purified ab137100 at 1:100 dilution (5.1 µg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) 1:200 (2.5 µg/ml). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) was used as the secondary antibody at 1:1000 (2 µg/ml) dilution. DAPI nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

Flow Cytometry analysis of SH-SY5Y (Human neuroblastoma epithelial cell) cells labeling Staufen/STAU1 with Purified ab137100 at 1:50 dilution (10 µg/ml) (red). Cells were fixed with 4% Paraformaldehyde. A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) secondary antibody was used at 1:2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

Anti-Staufen/STAU1 antibody [EPR7966] (ab137100) at 1/1000 dilution (Purified) + Rat heart lysates at 15 µg

Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Predicted band size: 63 kDa
Observed band size: 55,63 kDa why is the actual band size different from the predicted?

All lanes : Anti-Staufen/STAU1 antibody [EPR7966] (ab137100) at 1/1000 dilution (Purified)

Lane 1 : HepG2 (Human hepatocellular carcinoma epithelial cell) whole cell lysates
Lane 2 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysates
Lane 3 : Mouse brain lysates
Lane 4 : NIH/3T3 (Mouse embryonic fibroblast) whole cell lysates
Lane 5 : Neuro-2a (Mouse neuroblastoma neuroblast) whole cell lysates

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Predicted band size: 63 kDa
Observed band size: 55,63 kDa why is the actual band size different from the predicted?

All lanes : Anti-Staufen/STAU1 antibody [EPR7966] (ab137100) at 1/1000 dilution

Lane 1 : Jurkat cell lysate
Lane 2 : K562 cell lysate
Lane 3 : HepG2 cell lysate

Lysates/proteins at 10 µg per lane.

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

Predicted band size: 63 kDa
Observed band size: 55 kDa why is the actual band size different from the predicted?

This image was generated using the unpurified version of the product. 

Overlay histogram showing SHSY-5Y cells stained with ab137100 (red line). The cells were fixed with 4% paraformaldehyde (10 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 (ab137100, 1/10000 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. This antibody gave a positive signal in SHSY-5Y cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

This image was generated using the unpurified version of the product. 

Immunofluorescent analysis of HeLa cells labelling Staufen/STAU1 with ab137100 at 1/250 dilution.

This image was generated using the unpurified version of the product. 

Immunofluorescent analysis of HepG2 cells labelling Staufen/STAU1 with ab137100 at 1/250 dilution.

This image was generated using the unpurified version of the product.