Anti-c-Myc (phospho T58) antibody [EPR17923]

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-c-Myc (phospho T58) antibody [EPR17923] (AB185655)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa cells (Human epithelial cells from cervix adenocarcinoma) labeling c-Myc (phospho T58) with ab185655 at 1/250, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/500 (green).

Confocal image showing nuclear staining on HeLa cells. The staining decreased after blocking with phospho peptide (100μg/ml) overnight. The control peptide is a non-phospho peptide.

The nuclear counterstain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin antibody -Loading Control (ab7291) at 1/1000 dilution Goat Anti-Mouse IgG (AlexaFluor®594) preadsorbed (ab150120) at 1/500 (red).

The negative controls are as follows : -
-ve control 1 - ab185655 at 1/500 followed by ab150120 at 1/500.
-ve control 2 -ab7291 at 1/1000 followed by ab150077 at 1/500.

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-c-Myc (phospho T58) antibody [EPR17923] (AB185655)

Flow Cytometry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) treated with 200nM Calyculin A and 1uM Okadaic Acid for 60 min cells labeling c-Myc with purified ab185655 at 1/700 dilution (1 μg/mL) (Red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) secondary antibody was used at 1/2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

• WB

Supplier Data

Western blot - Anti-c-Myc (phospho T58) antibody [EPR17923] (AB185655)

Blocking and diluting buffer was 5% NFDM /TBST.

The strong band in Lane 1 of WB-2 compared to WB-1 is due to different lysate batches and a lower dilution factor (1 : 1000).

All lanes:

Western blot - Anti-c-Myc (phospho T58) antibody [EPR17923] (ab185655) at 1/1000 dilution

Lane 1:

HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 10 µg

Lane 2:

HeLa (Human epithelial cell line from cervix adenocarcinoma) treated with Lambda Phosphatase whole cell lysate at 10 µg

Secondary

All lanes:

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

Predicted band size: 48 kDa

Observed band size: 57 kDa

true

Exposure time: 3min

• WB

Supplier Data

Western blot - Anti-c-Myc (phospho T58) antibody [EPR17923] (AB185655)

Blocking and diluting buffer was 5% NFDM /TBST

All lanes:

Western blot - Anti-c-Myc (phospho T58) antibody [EPR17923] (ab185655) at 1/5000 dilution

Lane 1:

Untreated HeLa (Human epithelial cell line from cervix adenocarcinoma ) whole cell lysate at 10 µg

Lane 2:

HeLa (Human epithelial cell line from cervix adenocarcinoma ) treated with 200nM Calyculin A and 1uM Okadaic Acid for 60 minutes whole cell lysate at 10 µg

Secondary

All lanes:

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

Predicted band size: 48 kDa

Observed band size: 57 kDa

true

Exposure time: 3min

• Dot

Lab

Dot Blot - Anti-c-Myc (phospho T58) antibody [EPR17923] (AB185655)

Lane 1 : c-Myc (phospho T58).
Lane 2 : c-Myc (pT58) non-phospho peptide.
Lane 3 : c-Myc (pS62) phospho peptide.
Lane 4 : c-Myc (pS62) non-phospho peptide.

Dot blot analysis using ab185655 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 : 3 minutes.

• WB

CiteAb

Western blot - Anti-c-Myc (phospho T58) antibody [EPR17923] (AB185655)

c-Myc (phospho T58) western blot using anti-c-Myc (phospho T58) antibody [EPR17923] ab185655. Publication image and figure legend from Kazi, A., Xiang, S., et al., 2018, Nat Commun, PubMed 30514931.

ab185655 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab185655 please see the product overview.

SB blocks the phosphorylation of β-catenin S33/S37/T41 and T58 c-Myc. a Western blots of MiaPaCa2 and A549 tumor lysates from xenografts resected from mice treated with vehicle or SB. C1, C2, and C3 are tumors from mice treated with vehicle control. SB1, SB2, and SB3 are tumors from mice treated with SB. Experiment was done once. b Western blot of MiaPaCa2 and A549 cells treated with 1 µM SB for various lengths of time. Mia designates MiaPaCa2 lysates used as positive control. Experiment was done twice

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