Anti-SIKE1 antibody [EPR14692]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIKE1 antibody [EPR14692] (AB183509)

Immunohistochemical analysis of paraffin embedded Human kidney tissue labeling SIKE1 with ab183509 at 1/50.

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

• WB

Lab

Western blot - Anti-SIKE1 antibody [EPR14692] (AB183509)

Lanes 1 - 4 : Merged signal (red and green). Green - ab183509 observed at 25 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab183509 was shown to specifically react with SIKE1 when SIKE1 knockout samples were used. Wild-type and SIKE1 knockout samples were subjected to SDS-PAGE. ab183509 and ab8245 (loading control to GAPDH) were both diluted 1/10 000 and incubated overnight at 4°C. 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 1/10000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-SIKE1 antibody [EPR14692] (ab183509) at 1/10000 dilution

Lane 1:

Wild-type HAP1 cell lysate at 20 µg

Lane 2:

SIKE1 knockout HAP1 cell lysate at 20 µg

Lane 3:

HeLa cell lysate at 20 µg

Lane 4:

PC3 cell lysate at 20 µg

Predicted band size: 24 kDa

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• IP

Supplier Data

Immunoprecipitation - Anti-SIKE1 antibody [EPR14692] (AB183509)

Lysate from 293 cells was immunoprecipitated with ab183509 at a 1/50 dilution. For the subsequent blot, ab183509 used at a 1/10000 dilution with an anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at a 1/1500 dilution for the secondary. Blocking/ Dilution buffer : 5% NFDM/TBST. Lane 2 : TBS instead of lysate.

All lanes:

Immunoprecipitation - Anti-SIKE1 antibody [EPR14692] (ab183509)

Predicted band size: 24 kDa

Observed band size: 24 kDa

false

• WB

Supplier Data

Western blot - Anti-SIKE1 antibody [EPR14692] (AB183509)

Lanes 1-3 : Merged signal (red and green). Green - ab183509 observed at 25 kDa. Red - loading control ab8245 observed at 37 kDa.

ab183509 Rabbit monoclonal [EPR14692] to SIKE1 was shown to specifically react with Suppressor of IKBKE 1 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265933 (knockout cell lysate ab258188) was used. Wild-type and Suppressor of IKBKE 1 knockout samples were subjected to SDS-PAGE. ab183509 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 10000 dilution and 1 in 20000 dilution respectively. 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 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-SIKE1 antibody [EPR14692] (ab183509) at 1/10000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

SIKE1 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human SIKE1 knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-sike1-knockout-hela-cell-line-ab265933'>ab265933</a>)

Lane 3:

HEK-293 cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/20000 dilution

Predicted band size: 24 kDa

Observed band size: 25 kDa

false

• WB

Supplier Data

Western blot - Anti-SIKE1 antibody [EPR14692] (AB183509)

All lanes:

Western blot - Anti-SIKE1 antibody [EPR14692] (ab183509) at 1/20000 dilution

Lane 1:

SH-SY5Y cell lysate at 20 µg

Lane 2:

Hela cell lysate at 20 µg

Lane 3:

293 cell lysate at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated at 1/1000 dilution

Predicted band size: 24 kDa

Observed band size: 24 kDa

false