Anti-EGFR (phospho Y1086) antibody [Y39] - BSA and Azide free

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-EGFR (phospho Y1086) antibody [Y39] - BSA and Azide free (AB247225)

This data was developed using ab32086, the same antibody clone in a different buffer formulation.

Immunocytochemistry/ Immunofluorescence analysis of A431 (Human epidermoid carcinoma cell line) cells labeling EGFR (phospho Y1086) with ab32086 at 1/100, 3 μg/ml. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% tritonX-100. ab150077, a AlexaFluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1000, 2 μg/ml. Cells were counterstained with ab195889, anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594) at 1/200, 2.5 μg/ml. Nuclear stain was DAPI (blue).

The green staining on the membrane was increased in the EGF (100ng/ml, 10min) treated A431 cells when compared with A431 cells without treatment. After LP treatment, the green signaling was obviously decreased.

For the pan antibody, there was no great difference after EGF (100ng/ml, 10min) or EGF (100ng/ml, 10min) + LP treatment. The data showed mostly membranous staining.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-EGFR (phospho Y1086) antibody [Y39] - BSA and Azide free (AB247225)

This data was developed using ab32086, the same antibody clone in a different buffer formulation.Immunofluorescent analysis of EGFR (phospho Y1086) expression in A431 cell culture, using 1/100 ab32086.

• IP

Lab

Immunoprecipitation - Anti-EGFR (phospho Y1086) antibody [Y39] - BSA and Azide free (AB247225)

This data was developed using ab32086, the same antibody clone in a different buffer formulation.
Purified ab32086 at 1/20 dilution (1μg) immunoprecipitating EGFR in A431 treated with 100ng/mL EGF for 30min whole cell lysate.
Lane 1 (input) : A431 (Human epidermoid carcinoma epithelial cell) treated with 100ng/mL EGF for 30min whole cell lysate 10μg
Lane 2 (+) : ab32086 + A431 treated with 100ng/mL EGF for 30min whole cell lysate.
Lane 3 (-) : Rabbit monoclonal IgG (ab172730) instead of ab32086 in A431 treated with 100ng/mL EGF for 30min whole cell lysate.
VeriBlot for IP Detection Reagent (HRP) (ab131366) (1/1000 dilution) was used for Western blotting.
Blocking Buffer and concentration : 5% NFDM/TBST.
Diluting buffer and concentration : 5% NFDM/TBST.
Observed band size : 175 kDa

All lanes:

Immunoprecipitation - Anti-EGFR (phospho Y1086) antibody [Y39] (<a href='/en-us/products/primary-antibodies/egfr-phospho-y1086-antibody-y39-ab32086'>ab32086</a>)

Predicted band size: 129 kDa,134 kDa

false

• WB

Lab

Western blot - Anti-EGFR (phospho Y1086) antibody [Y39] - BSA and Azide free (AB247225)

This data was developed using ab32086, the same antibody clone in a different buffer formulation.

Blocking and dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-EGFR (phospho Y1086) antibody [Y39] (<a href='/en-us/products/primary-antibodies/egfr-phospho-y1086-antibody-y39-ab32086'>ab32086</a>) at 1/3000 dilution

Lane 1:

Untreated A431 (Human epidermoid carcinoma cell line) whole cell lysates at 15 µg

Lane 2:

A431 (Human epidermoid carcinoma cell line) treated with 100 ng/ml EGF for 10 minutes whole cell lysates at 15 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: 134 kDa

Observed band size: 170 kDa

false

Exposure time: 10s

• WB

Unknown

Western blot - Anti-EGFR (phospho Y1086) antibody [Y39] - BSA and Azide free (AB247225)

This data was developed using ab32086, the same antibody clone in a different buffer formulation.

All lanes:

Western blot - Anti-EGFR (phospho Y1086) antibody [Y39] (<a href='/en-us/products/primary-antibodies/egfr-phospho-y1086-antibody-y39-ab32086'>ab32086</a>) at 1/10000 dilution

Lane 1:

Untreated A431 cell lysate

Lane 2:

A431 cell lysate, treated with EGF

Predicted band size: 134 kDa

Observed band size: 150 kDa

false

• Dot

Unknown

Dot Blot - Anti-EGFR (phospho Y1086) antibody [Y39] - BSA and Azide free (AB247225)

This data was developed using ab32086, the same antibody clone in a different buffer formulation.

Dot blot analysis of EGFR (phospho Y1086) phospho peptide (Lane 1), EGFR non-phospho peptide (Lane 2), labeling EGFR (phospho Y1086) with ab32068 at a dilution of 1/1000. Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) was used as the secondary antibody at a dilution of 1/100000.

Blocking and dilution buffer : 5% NFDM/TBST.

Exposure time : 5 seconds.