Western blot - anti-c-Myc antibody [9E10] (ab32)

Anti-c-Myc antibody [9E10] - ChIP Grade (ab32) at 1 µg/ml + E. coli Positive Control (Escherichia coli ) Whole Cell Lysate (ab5395) at 10 µg

Secondary
Goat polyclonal to Mouse IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution
developed using the ECL technique

Performed under reducing conditions.

Predicted band size : 41 kDa
Observed band size : 45 kDa (why is the actual band size different from the predicted?)


Exposure time : 1 minute

Flow Cytometry-Anti-c-Myc antibody [9E10] - ChIP Grade(ab32)

Overlay histogram showing Jurkat cells stained with ab32 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab32, 1µg/1x10⁶ cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed.

Western blot - c-Myc antibody [9E10] (ab32)

Predicted band size : 41 kDa


Phosphorylation of Cdc15 changes during the cell cycle. Exponentially growing cells (cyc) of CDC15-MYC9 (W1114) were arrested in G1 with a factor pheromone (a) and released into fresh medium at 258C. Cells were harvested at the indicated times, the percentage of divided nuclei was determined by DAPI staining of fixed cells,and proteins were analyzed by Western blotting with 9E10 (ab32).

Menssen R et al., (2001) Curr Biol. Mar 6;11(5):345-50.

Immunocytochemistry/ Immunofluorescence - c-Myc antibody [9E10] (ab32)

ICC/IF image of ab32 stained human Hek293 cells. The cells were methanol fixed (10 min), permabilised in TBS-T (20 min) and incubated with the antibody (ab32, 5µg/ml) for 1h at room temperature. 1%BSA / 10% normal serum / 0.3M glycine was used to quench autofluorescence and block non-specific protein-protein interactions. The secondary antibody (green) was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red). DAPI was used to stain the cell nuclei (blue).