Anti-MEK1 (phospho T286) antibody [EPR27226-8] - BSA and Azide free

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-MEK1 (phospho T286) antibody [EPR27226-8] - BSA and Azide free (AB307510)

This data was produced using ab307509, the same clone but in a different formulation. Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized MAP2K1 KO HAP1 (MAP2K1 knockout human chronic myelogenous leukemia near-haploid cell) cells labelling MEK1 (phospho T286) with ab307509 at 1/1000 (0.48 ug/ml) followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (2 ug/ml) (Green). Confocal showing cytoplasmic and nuclear staining in wildtype HAP1 cells in M phase and no staining in MAP2K1 knockout HAP1 cells. The image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (2.5 ug/ml) (Red). The nuclear counterstain was DAPI (Blue). Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (2 ug/ml).

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-MEK1 (phospho T286) antibody [EPR27226-8] - BSA and Azide free (AB307510)

This data was produced using ab307509, the same clone but in a different formulation. Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NIH/3T3 (mouse embryonic fibroblast) cells labelling MEK1 (phospho T286) with ab307509 at 1/1000 (0.48 ug/ml) followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (2 ug/ml) (Green). Confocal showing cytoplasmic and nuclear staining in NIH/3T3 cells in M phase. The image was taken with a confocal microscope (Leica-Microsystems, TCS SP8). ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (2.5 ug/ml) (Red). The nuclear counterstain was DAPI (Blue). Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution (2 ug/ml).

• WB

Supplier Data

Western blot - Anti-MEK1 (phospho T286) antibody [EPR27226-8] - BSA and Azide free (AB307510)

This data was produced using ab307509, the same clone but in a different formulation. Blocking and diluting buffer and concentration : 1% BSA/TBST. The identity of the higher MW bands between 100kDa and 200kDa are unknown. Lysates were freshly made and used for Western blotting immediately to minimize protein degradation. In Western blot, anti-GAPDH antibody (ab181602) loading control staining at 1/200000 dilution. In Western blot, anti-MEK1 antibody (ab32576) loading control staining at 1/1000 dilution. Exposure time : 26 seconds.

All lanes:

Western blot - Anti-MEK1 (phospho T286) antibody [EPR27226-8] (<a href='/en-us/products/primary-antibodies/mek1-phospho-t286-antibody-epr27226-8-ab307509'>ab307509</a>) at 1/1000 dilution

Lane 1:

Untreated HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate (untreated membrane) at 20 µg

Lane 2:

HeLa treated with 100ng/ml nocodazole for 24h whole cell lysate (untreated membrane) at 20 µg

Lane 3:

HeLa treated with 100ng/ml nocodazole for 24h whole cell lysate (phosphatase treated membrane at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 43 kDa

Observed band size: 43 kDa

false

Exposure time: 26s

• WB

Supplier Data

Western blot - Anti-MEK1 (phospho T286) antibody [EPR27226-8] - BSA and Azide free (AB307510)

This data was produced using ab307509, the same clone but in a different formulation. Blocking and diluting buffer and concentration : 1% BSA/TBST. The identity of the higher MW bands between 100kDa and 200kDa are unknown. Lysates were freshly made and used for Western blotting immediately to minimize protein degradation. In Western blot, anti-GAPDH antibody (ab181602) loading control staining at 1/200000 dilution. In Western blot, anti-MEK1 antibody (ab32576) loading control staining at 1/1000 dilution. Exposure time : 15 seconds.

All lanes:

Western blot - Anti-MEK1 (phospho T286) antibody [EPR27226-8] (<a href='/en-us/products/primary-antibodies/mek1-phospho-t286-antibody-epr27226-8-ab307509'>ab307509</a>) at 1/1000 dilution

Lane 1:

Untreated wild-type HAP1(human chronic myelogenous leukemia near-haploid cell line) whole cell lysate (untreated membrane), at 20 µg

Lane 2:

Wild-type HAP1 treated with 100ng/ml nocodazole for 24h whole cell lysate (untreated membrane) at 20 µg

Lane 3:

untreated MEK1 knockout HAP1 whole cell lysate at 20 µg

Lane 4:

MEK1 knockout HAP1 treated with 100ng/ml nocodazole for 24h whole cell lysate at 20 µg

Lane 5:

Untreated wild-type HAP1 whole cell lysate (phosphatase treated membrane), at 20 µg

Lane 6:

Wild-type HAP1 treated with 100ng/ml nocodazole for 24h whole cell lysate (phosphatase treated membrane) at 20 µg

Lane 7:

Untreated MEK1 knockout HAP1 whole cell lysate (phosphatase treated membrane) at 20 µg

Lane 8:

MEK1 knockout HAP1 treated with 100ng/ml nocodazole for 24h whole cell lysate (phosphatase treated membrane) at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 43 kDa

Observed band size: 43 kDa

false

Exposure time: 15s

• WB

Supplier Data

Western blot - Anti-MEK1 (phospho T286) antibody [EPR27226-8] - BSA and Azide free (AB307510)

This data was produced using ab307509, the same clone but in a different formulation.

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

Lysates were freshly made and used for Western blotting immediately to minimize protein degradation.

This blot was developed using a high sensitivity ECL substrate. The high-sensitivity ECL substrate used allows for the detection of proteins in the mid-femtogram range.

In Western blot, anti-GAPDH antibody (ab181602) loading control staining at 1/200000 dilution.

In Western blot, anti-MEK1+MEK2 antibody (ab178876) loading control staining at 1/1000 dilution.

All lanes:

Western blot - Anti-MEK1 (phospho T286) antibody [EPR27226-8] (<a href='/en-us/products/primary-antibodies/mek1-phospho-t286-antibody-epr27226-8-ab307509'>ab307509</a>) at 1/1000 dilution

Lane 1:

Untreated NIH/3T3 (mouse embryonic fibroblast) whole cell lysate (untreated membrane) at 20 µg

Lane 2:

NIH/3T3 treated with 100ng/ml nocodazole for 24h whole cell lysate (untreated membrane) at 20 µg

Lane 3:

NIH/3T3 treated with 100ng/ml nocodazole for 24h whole cell lysate (phosphatase treated membrane) at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Predicted band size: 43 kDa

Observed band size: 43 kDa

true

Exposure time: 82s

• Dot

Supplier Data

Dot Blot - Anti-MEK1 (phospho T286) antibody [EPR27226-8] - BSA and Azide free (AB307510)

This data was produced using ab307509, the same clone but in a different formulation.

Dot blot analysis of MEK1 (phospho T286) using ab307509 at 1 : 1000 (0.48 ug/ml) followed by a Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1 : 100,000 dilution.

Exposure time : 18 seconds.

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

Lane 1 : MEK1 (phospho T286) peptide a

Lane 2 : MEK1 (phospho T286) peptide b

Lane 3 : MEK1 non-phospho peptide

All lanes:

Dot Blot - Anti-MEK1 (phospho T286) antibody [EPR27226-8] (<a href='/en-us/products/primary-antibodies/mek1-phospho-t286-antibody-epr27226-8-ab307509'>ab307509</a>) at 1/1000 dilution

Lane 1:

MEK1 (phospho T286) peptide a

Lane 2:

MEK1 (phospho T286) peptide b

Lane 3:

MEK1 non-phospho peptide

Secondary

All lanes:

Dot Blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

false