All lanes : Anti-Nucleophosmin (phospho T199) antibody [EP1857Y] (ab81551) at 1/2500 dilution

Lane 1 : lysate from untreated HeLa cells
Lane 2 : lysate from HeLa cells treated with CA

Lysates/proteins at 10 µg per lane.

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

Predicted band size: 33 kDa
Observed band size: 33 kDa

Immunohistochemistry analysis of paraffin-embedded Human lymphoma tissue, using 1/100 ab81551.

Perform heat mediated antigen retrieval with EDTA buffer pH 9 before commencing with IHC staining protocol.

Immunofluorescent staing of HeLa cells using 1/100 ab81551.

ab81551 staining Nucleophosmin (phospho T199) in human HeLa cells by Immunocytochemistry/ Immunofluorescence. Cells were fixed with formaldehyde, permeabilized with 0.1% Triton x100 in PBS and blocking with 5% serum was performed for 30 minutes at 23⁰C. Samples were incubated with primary antibody (1/150 in PBS) for 1 hour at 23°C. An Alexa Fluor^®488-conjugated goat polyclonal to rabbit IgG was used at dilution at 1/100 as secondary antibody.

See Abreview

Overlay histogram showing HeLa cells stained with ab76539 (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 (ab76539, 1/100 dilution) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1µg/1x10⁶ cells) used under the same conditions. Unlabelled sample (blue line). Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.