Anti-USP22 antibody [EPR4352(2)] - BSA and Azide free

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-USP22 antibody [EPR4352(2)] - BSA and Azide free (AB223341)

This data was developed using ab109435, the same antibody clone in a different buffer formulation.

Intracellular flow cytometric analysis of permeabilized HeLa cells using 1/10 ab109435 (red) or a rabbit IgG (negative) (green).

• WB

Lab

Western blot - Anti-USP22 antibody [EPR4352(2)] - BSA and Azide free (AB223341)

False colour image of Western blot : Anti-USP22 antibody [EPR4352(2)] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab109435 was shown to bind specifically to USP22. A band was observed at 59 kDa in wild-type HeLa cell lysates with no signal observed at this size in usp22 knockout cell line ab264888 (knockout cell lysate ab257789). To generate this image, wild-type and usp22 knockout HeLa cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged.Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

All lanes:

Western blot - Anti-USP22 antibody [EPR4352(2)] (<a href='/en-us/products/primary-antibodies/usp22-antibody-epr43522-ab109435'>ab109435</a>) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human USP22 knockout HeLa cell lysate (<a href='/en-us/products/cell-lysates/human-usp22-knockout-hela-cell-lysate-ab257789'>ab257789</a>) at 20 µg

Predicted band size: 60 kDa

Observed band size: 59 kDa

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• WB

Lab

Western blot - Anti-USP22 antibody [EPR4352(2)] - BSA and Azide free (AB223341)

This data was developed using ab109435, the same antibody clone in a different buffer formulation.

Lane 1 : Wild-type HAP1 cell lysate (20 μg)

Lane 2 : USP22 knockout HAP1 cell lysate (20 μg)

Lane 3 : HeLa cell lysate (20 μg)

Lane 4 : HepG2 cell lysate (20 μg)

Lanes 1 - 4 : Merged signal (red and green). Green - ab109435 observed at 60 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab109435 was shown to specifically react with USP22 in wild-type cells along with additional cross-reactive bands. The band was not seen in USP22 knockout HAP1 cells. Wild-type and USP22 knockout samples were subjected to SDS-PAGE. ab109435 and ab8245 (loading control to GAPDH) were diluted at 1/1000 and 1/10000 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 h at room temperature before imaging.

All lanes:

Western blot - Anti-USP22 antibody [EPR4352(2)] (<a href='/en-us/products/primary-antibodies/usp22-antibody-epr43522-ab109435'>ab109435</a>)

Predicted band size: 60 kDa

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• WB

Unknown

Western blot - Anti-USP22 antibody [EPR4352(2)] - BSA and Azide free (AB223341)

This data was developed using ab109435, the same antibody clone in a different buffer formulation.

All lanes:

Western blot - Anti-USP22 antibody [EPR4352(2)] (<a href='/en-us/products/primary-antibodies/usp22-antibody-epr43522-ab109435'>ab109435</a>) at 1/1000 dilution

Lane 1:

HeLa cell lysate at 10 µg

Lane 2:

Human placenta lysate at 10 µg

Lane 3:

HT-1376 lysate at 10 µg

Predicted band size: 60 kDa

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• OI-RD Scanning

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OI-RD Scanning - Anti-USP22 antibody [EPR4352(2)] - BSA and Azide free (AB223341)

We have systematically measured KD (the equilibrium dissociation constant between the antibody and its antigen), of more than 840 recombinant antibodies to assess not only their individual KD values but also to see the average affinity of antibody. Based on the comparison with published literature values for mouse monoclonal antibodies, Recombinant antibodies appear to be on average 1-2 order of magnitude higher affinity.