Anti-RSK4 antibody [EP1982Y]

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-RSK4 antibody [EP1982Y] (AB76117)

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

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-RSK4 antibody [EP1982Y] (AB76117)

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.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-RSK4 antibody [EP1982Y] (AB76117)

Immunohistochemical staining of paraffin embedded human kidney with purified ab76117 at a working dilution of 1/250. The secondary antibody used is HRP goat anti-rabbit IgG H&L (ab97051) at 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-RSK4 antibody [EP1982Y] (AB76117)

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.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-RSK4 antibody [EP1982Y] (AB76117)

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).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-RSK4 antibody [EP1982Y] (AB76117)

Immunohistochemical staining of paraffin embedded rat kidney with purified ab76117 at a working dilution of 1/250. The secondary antibody used is HRP goat anti-rabbit IgG H&L (ab97051) at 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-RSK4 antibody [EP1982Y] (AB76117)

Immunohistochemical staining of paraffin embedded mouse cerebral cortex with purified ab76117 at a working dilution of 1/250. The secondary antibody used is HRP goat anti-rabbit IgG H&L (ab97051) at 1/500. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

• WB

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Western blot - Anti-RSK4 antibody [EP1982Y] (AB76117)

All lanes:

Western blot - Anti-RSK4 antibody [EP1982Y] (ab76117) at 1/500 dilution

All lanes:

SH-SY5Y cell lysate at 10 µg

Secondary

All lanes:

HRP labelled goat anti-rabbit at 1/2000 dilution

Predicted band size: 84 kDa

Observed band size: 90 kDa

false

• WB

Lab

Western blot - Anti-RSK4 antibody [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.

All lanes:

Western blot - Anti-RSK4 antibody [EP1982Y] (ab76117)

Predicted band size: 39 kDa,84 kDa

false

• WB

Lab

Western blot - Anti-RSK4 antibody [EP1982Y] (AB76117)

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

All lanes:

Western blot - Anti-RSK4 antibody [EP1982Y] (ab76117) at 1/1000 dilution

Lane 1:

SH-SY5Y cell lysate at 20 µg

Lane 2:

Caco-2 cell lysate at 20 µg

Secondary

All lanes:

HRP goat anti-rabbit IgG (H+L) at 1/1000 dilution

Predicted band size: 84 kDa

Observed band size: 90 kDa

false