重组Anti-MEK3抗体[EPR17345-104]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MEK3 antibody [EPR17345-104] (AB195037)

Immunohistochemical analysis of paraffin-embedded Human skeletal muscle tissue labeling MEK3 with ab195037 at 1/500 dilution followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) secondary at 1/500 dilution. Cytoplasmic and nucleus staining on Human skeletal muscle is observed. Counter stained with Hematoxylin.

Negative control : Using PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

1 : Ben-Levy R,et.al.(1998) Nuclear export of the stress-activated protein kinase p38 mediated by its substrate MAPKAP kinase-2. Curr Biol, 8(19) : 1049-1057.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MEK3 antibody [EPR17345-104] (AB195037)

Immunohistochemical analysis of paraffin-embedded Human lung adenocarcinoma tissue labeling MEK3 with ab195037 at 1/500 dilution followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) secondary at 1/500 dilution. Cytoplasmic and nuclear staining on cancer cells of Human lung adenocarcinoma is observed. Counter stained with Hematoxylin.

Negative control : Using PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IP

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Immunoprecipitation - Anti-MEK3 antibody [EPR17345-104] (AB195037)

MEK3 was immunoprecipitated from 1mg of HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell extract with ab195037 at 1/90 dilution. Western blot was performed from the immunoprecipitate using ab195037 at 1/1000 dilution. Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG, was used as secondary antibody at 1/1500 dilution. Lane 1 : HeLa whole cell extract. Lane 2 : PBS instead of HeLa whole cell extract.

Blocking and dilution buffer and concentration : 5% NFDM/TBST.

Based on the sequence analysis, ab195037 recognizes 3 isoforms of MEK3 ranging from 36kDa to 40KDa. The multi-bands should be MEK3 isoforms.

All lanes:

Immunoprecipitation - Anti-MEK3 antibody [EPR17345-104] (ab195037)

Predicted band size: 39 kDa

false

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MEK3 antibody [EPR17345-104] (AB195037)

Immunohistochemical analysis of paraffin-embedded Mouse skeletal muscle tissue labeling MEK3 with ab195037 at 1/500 dilution followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) secondary at 1/500 dilution. Cytoplasmic and nuclear staining on mouse skeletal muscle is observed. Counter stained with Hematoxylin.

Negative control : Using PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-MEK3 antibody [EPR17345-104] (AB195037)

Immunohistochemical analysis of paraffin-embedded Rat skeletal muscle tissue labeling MEK3 with ab195037 at 1/500 dilution followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) secondary at 1/500 dilution. Cytoplasmic and nuclear staining on rat skeletal muscle is observed. Counter stained with Hematoxylin.

Negative control : Using PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-MEK3 antibody [EPR17345-104] (AB195037)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (Mouse embyro fibroblast cells) cells labeling MEK3 with ab195037 at 1/250 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/500 dilution (green). Confocal image showing both nuclear and cytoplasmic staining on NIH/3T3 cell line. The nuclear counter stain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 dilution (red).
The negative controls are as follows :
-ve control 1 : ab195037 at 1/250 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/500 dilution.
-ve control 2 : ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/500 dilution.

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-MEK3 antibody [EPR17345-104] (AB195037)

Intracellular Flow Cytometry analysis of NIH/3T3 (mouse embryo) labelling MEK3 with purified ab195037 at 1/1000 (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. Alexa Fluor^® 488 goat anti-rabbit IgG (1/2000) was used as the secondary antibody. Black - Isotype control, rabbit monoclonal IgG. Blue - Unlabelled control, cells without incubation with primary and secondary antibodies.

• WB

Supplier Data

Western blot - Anti-MEK3 antibody [EPR17345-104] (AB195037)

Blocking/Dilution buffer : 5% NFDM/TBST.

Based on the sequence analysis, ab195037 recognizes 3 isoforms of MEK3 ranging from 36kDa to 40KDa. The multi-bands should be MEK3 isoforms.

All lanes:

Western blot - Anti-MEK3 antibody [EPR17345-104] (ab195037) at 1/5000 dilution

All lanes:

Human skeletal muscle lysates at 10 µg

Secondary

All lanes:

Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution

Predicted band size: 39 kDa

Observed band size: 34 kDa,39 kDa

false

• WB

Lab

Western blot - Anti-MEK3 antibody [EPR17345-104] (AB195037)

Lane 1 : Wild-type HAP1 cell lysate (20 μg)
Lane 2 : MEK3 knockout HAP1 cell lysate (20 μg)
Lane 3 : HeLa cell lysate (20 μg)
Lane 4 : Jurkat cell lysate (20 μg)
Lanes 1 - 4 : Merged signal (red and green). Green - ab195037 observed at 40 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab195037 was shown to specifically react with MEK3 when MEK3 knockout samples were used. Wild-type and MEK3 knockout samples were subjected to SDS-PAGE. ab195037 and ab8245 (loading control to GAPDH) were both diluted to 1/5000 and 1/10000 respectively 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-MEK3 antibody [EPR17345-104] (ab195037)

Predicted band size: 39 kDa

false

• WB

Supplier Data

Western blot - Anti-MEK3 antibody [EPR17345-104] (AB195037)

Blocking/Dilution buffer : 5% NFDM/TBST.

Based on the sequence analysis, ab195037 recognizes 3 isoforms of MEK3 ranging from 36kDa to 40KDa. The multi-bands should be MEK3 isoforms.

All lanes:

Western blot - Anti-MEK3 antibody [EPR17345-104] (ab195037) at 1/50000 dilution

All lanes:

HepG2 (Human liver hepatocellular carcinoma) whole cell lysates at 10 µg

Secondary

All lanes:

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

Predicted band size: 39 kDa

Observed band size: 34 kDa,39 kDa

false

• WB

Supplier Data

Western blot - Anti-MEK3 antibody [EPR17345-104] (AB195037)

Blocking/Dilution buffer : 5% NFDM/TBST.

Based on the sequence analysis, ab195037 recognizes 3 isoforms of MEK3 ranging from 36kDa to 40KDa. The multi-bands should be MEK3 isoforms.

All lanes:

Western blot - Anti-MEK3 antibody [EPR17345-104] (ab195037) at 1/10000 dilution

Lane 1:

HeLa (Human epithelial cells from cervix adenocarcinoma) whole cell lysates at 20 µg

Lane 2:

MCF7 (Human breast adenocarcinoma cell line) whole cell lysates at 20 µg

Secondary

All lanes:

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

Predicted band size: 39 kDa

Observed band size: 34 kDa,39 kDa

false

• WB

Supplier Data

Western blot - Anti-MEK3 antibody [EPR17345-104] (AB195037)

Blocking/Dilution buffer : 5% NFDM/TBST.

Based on the sequence analysis, ab195037 recognizes 3 isoforms of MEK3 ranging from 36kDa to 40KDa. The multi-bands should be MEK3 isoforms.

All lanes:

Western blot - Anti-MEK3 antibody [EPR17345-104] (ab195037) at 1/10000 dilution

Lane 1:

Mouse spleen lysates at 10 µg

Lane 2:

Rat spleen lysates at 10 µg

Secondary

All lanes:

Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution

Predicted band size: 39 kDa

false