Anti-IKKi/IKKe antibody

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-IKKi/IKKe antibody (AB7891)

Immunocytochemistry/Immunofluorecense analysis of HeLa cells with ab7891 at 20μg/ml.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IKKi/IKKe antibody (AB7891)

ab7891 at 10μg/ml staining IKK epsilon in human pancreatic tissue by IHC.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-IKKi/IKKe antibody (AB7891)

ICC/IF image of ab7891 stained HeLa cells. The cells were 100% methanol fixed (5 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab7891, 1μg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor^® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor^® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43μM.

• WB

Supplier Data

Western blot - Anti-IKKi/IKKe antibody (AB7891)

All lanes:

Western blot - Anti-IKKi/IKKe antibody (ab7891) at 1 µg/mL

Lane 1:

Untreated Raw264.7 cell lysates

Lane 2:

Raw264.7 cell lysates treated with LPS (0.3ug/ml) for 3 hrs

Lane 3:

Raw264.7 cell lysates treated with LPS (0.3ug/ml) for 6 hrs

Secondary

All lanes:

Goat anti-rabbit IgG HRP conjugate at 1/10000 dilution

Predicted band size: 80 kDa

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• WB

Unknown

Western blot - Anti-IKKi/IKKe antibody (AB7891)

All lanes:

Western blot - Anti-IKKi/IKKe antibody (ab7891) at 1 µg/mL

All lanes:

Jurkat whole cell lysate

Predicted band size: 80 kDa

Observed band size: 80 kDa

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• WB

CiteAb

Western blot - Anti-IKKi/IKKe antibody (AB7891)

Western Blotting using Anti-IKKi/IKKe antibody, ab7891. Publication image from Zhao, L. et al., 2020, Theranostics, 32104508. Legend direct from paper.

IKKε directly phosphorylates kindlin-2 at S159. (A) Slot blot analysis of the anti-phospho-kindlin-2-S159 antibody. The antibody was probed against an un-modified kindlin-2 peptide and phosphorylated peptides. (B) HCT116 cells were transfected with the indicated plasmids and whole cell extracts were immunoprecipitated (IP) with FLAG-conjugated M2 beads and then analyzed by immunoblotting (IB). (C)HCT116 cells were treated with the inhibitor IKK-3 at the indicated concentration for 24h and analyzed by immunoblotting. (D and E) HCT116 cells were transfected with the indicated siRNAs targeting IKKα, IKKβ, IKKε or TBK1. Cell lysates were analyzed by immunoblotting. (F) HCT116 cells were transfected with FLAG-tagged kindlin-2 and GFP-tagged IKKε for 48 h. Total cell lysates were immunoprecipitated using FLAG-conjugated M2 beads and analyzed by immunoblotting. (G) IKKε was immunoprecipitated using anti-IKKε antibody or the control antibody IgG from lysates of HCT116 cells. (H) HCT116 cells were transfected with the indicated plasmids and the whole cell extracts were prepared and analyzed by Co-IP assay flowed by immunoblotting with the indicated antibodies. (I) GST-tagged wild-type kindlin-2 or kindlin-2 (S159A) were incubated without IKKε (-), or with purified wild-type IKKε (WT) or mutant IKKε (K38A) in kinase buffer and resolved by SDS-PAGE, followed by immunoblotting analysis with the indicated antibodies.

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• WB

CiteAb

Western blot - Anti-IKKi/IKKe antibody (AB7891)

Western Blotting using Anti-IKKi/IKKe antibody, ab7891. Publication image from Zhao, L. et al., 2020, Theranostics, 32104508. Legend direct from paper.

IKKε directly phosphorylates kindlin-2 at S159. (A) Slot blot analysis of the anti-phospho-kindlin-2-S159 antibody. The antibody was probed against an un-modified kindlin-2 peptide and phosphorylated peptides. (B) HCT116 cells were transfected with the indicated plasmids and whole cell extracts were immunoprecipitated (IP) with FLAG-conjugated M2 beads and then analyzed by immunoblotting (IB). (C)HCT116 cells were treated with the inhibitor IKK-3 at the indicated concentration for 24h and analyzed by immunoblotting. (D and E) HCT116 cells were transfected with the indicated siRNAs targeting IKKα, IKKβ, IKKε or TBK1. Cell lysates were analyzed by immunoblotting. (F) HCT116 cells were transfected with FLAG-tagged kindlin-2 and GFP-tagged IKKε for 48 h. Total cell lysates were immunoprecipitated using FLAG-conjugated M2 beads and analyzed by immunoblotting. (G) IKKε was immunoprecipitated using anti-IKKε antibody or the control antibody IgG from lysates of HCT116 cells. (H) HCT116 cells were transfected with the indicated plasmids and the whole cell extracts were prepared and analyzed by Co-IP assay flowed by immunoblotting with the indicated antibodies. (I) GST-tagged wild-type kindlin-2 or kindlin-2 (S159A) were incubated without IKKε (-), or with purified wild-type IKKε (WT) or mutant IKKε (K38A) in kinase buffer and resolved by SDS-PAGE, followed by immunoblotting analysis with the indicated antibodies.

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