Anti-NEK7 antibody [EPR4901] - BSA and Azide free

• Flow Cyt (Intra)

Lab

Flow Cytometry (Intracellular) - Anti-NEK7 antibody [EPR4901] - BSA and Azide free (AB247860)

Flow cytometry overlay histogram showing left wild-type HAP1 WT and right NEK7 knockout HAP1 WT stained with ab247860 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilised with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS containing 10% normal goat serum to block non-specific protein-protein interaction followed by the antibody (ab109433) (1x 10⁶ in 100μl at 0.2 μg/ml (1/3135 dilution)) for 30min at 22°C.

The secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed was incubated at 1/4000 dilution for 30min at 22°C

Isotype control antibody Recombinant Rabbit IgG, monoclonal [EPR25A] was used at the same concentration and conditions as the primary antibody (black line). Unlabelled sample was also used as a control (blue line).

Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.

PFA fixation is not recommended.

• WB

Unknown

Western blot - Anti-NEK7 antibody [EPR4901] - BSA and Azide free (AB247860)

This data was developed using ab109433, the same antibody clone in a different buffer formulation.

All lanes:

Western blot - Anti-NEK7 antibody [EPR4901] (<a href='/en-us/products/primary-antibodies/nek7-antibody-epr4901-ab109433'>ab109433</a>) at 1/10000 dilution

Lane 1:

Human brain lysate at 10 µg

Lane 2:

HeLa cell lysate at 10 µg

Lane 3:

A549 cell lysate at 10 µg

Lane 4:

HepG2 cell lysate at 10 µg

Lane 5:

Fetal liver lysate at 10 µg

Lane 6:

Jurkat cell lysate at 10 µg

Predicted band size: 35 kDa

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• WB

Lab

Western blot - Anti-NEK7 antibody [EPR4901] - BSA and Azide free (AB247860)

This data was developed using ab109433, the same antibody clone in a different buffer formulation.

Lane 1 : Wild-type HAP1 cell lysate (40 μg)

Lane 2 : NEK7 knockout HAP1 cell lysate (40 μg)

Lane 3 : A549 cell lysate (20 μg)

Lane 4 : Jurkat cell lysate (20 μg)

Lanes 1 - 4 : Merged signal (red and green). Green - ab109433 observed at 35 kDa. Red - loading control, ab18058, observed at 124 kDa.

ab109433 was shown to specifically react with NEK7 when NEK7 knockout samples were used. Wild-type and NEK7 knockout samples were subjected to SDS-PAGE. ab109433 and ab18058 (loading control to Vinculin) were diluted at 1/1000 and 1/10000 dilution 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-NEK7 antibody [EPR4901] (<a href='/en-us/products/primary-antibodies/nek7-antibody-epr4901-ab109433'>ab109433</a>)

Predicted band size: 35 kDa

false