Anti-RSK4 抗体 [EP1982Y] (ab76117)

Lane 1: Wild-type HAP1 cell lysate (40 µg)
Lane 2: RSK4 knockout HAP1 cell lysate (40 µg)
Lane 3: MCF7 cell lysate (20 µg)
Lane 4: HEK293 cell lysate (20 µg)
Lanes 1 - 4: Merged signal (red and green). Green - ab76117 observed at 90 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab76117 was shown to recognize RSK4 when RSK4 knockout samples were used, along with additional cross-reactive bands. Wild-type and RSK4 knockout samples were subjected to SDS-PAGE. Ab76117 and ab8245 (loading control to GAPDH) were diluted at 1/250 and 1/10,000 dilution respectively and incubated overnight at 4C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10,000 dilution for 1 hour at room temperature before imaging.

Blocking buffer: 5% NFDM/TBST
Dilution buffer: 5% NFDM/TBST

Immunofluorescence staining of HEK293 cells with purified ab76117 at a working dilution of 1/1000, counter-stained with DAPI. The secondary antibody was Alexa Fluor^® 488 goat anti-rabbit (ab150077), used at a dilution of 1/1000. ab7291, a mouse anti-tubulin antibody (1/1000), was used to stain tubulin along with ab150120 (Alexa Fluor^® 594 goat anti-mouse, 1/1000), shown in the top right hand panel. The cells were fixed in 4% PFA and permeabilized using 0.1% Triton X 100. The negative controls are shown in bottom middle and right hand panels - for negative control 1, purified ab76117 was used at a dilution of 1/500 followed by an Alexa Fluor^® 594 goat anti-mouse antibody (ab150120) at a dilution of 1/500. For negative control 2, ab7291 (mouse anti-tubulin) was used at a dilution of 1/500 followed by an Alexa Fluor^® 488 goat anti-rabbit antibody (ab150077) at a dilution of 1/400.

Overlay histogram showing Neuro-2a cells fixed in 2% PFA and stained with purified ab76117 at a dilution of 1 in 70 (red line). The secondary antibody used was FITC goat anti-rabbit at a dilution of 1 in 500. Rabbit monoclonal IgG was used as an isotype control (black line) and cells incubated in the absence of both primary and secondary antibody were used as a negative control (blue line).

Unpurified ab76117, at 1/50 dilution, staining RSK4 in paraffin-embedded human brain glioma tissue.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Immunofluorescent staining of SH-Sy5y cells using unpurified ab76117 at 1/100 dilution.