Anti-USP22 antibody [EPR18945]

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-USP22 antibody [EPR18945] (AB195289)

Immunohistochemical analysis of paraffin-embedded Human cerebrum tissue labeling USP22 with ab195289 at 1/100 followed by a Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) Ready to use. Nuclear staining on human cerebrum. The section was incubated with ab195289 for 30 mins at room temperature.The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) Ready to use.

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-USP22 antibody [EPR18945] (AB195289)

Immunohistochemical analysis of paraffin-embedded Human breast carcinoma tissue labeling USP22 with ab195289 at 1/100 followed by a Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) at Ready to use dilution. Nuclear staining on human breast carcinoma. The section was incubated with ab195289 for 30 mins at room temperature.The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) Ready to use.

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins

• IP

Supplier Data

Immunoprecipitation - Anti-USP22 antibody [EPR18945] (AB195289)

USP22 was immunoprecipitated from 0.35 mg of HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab195289 at 1/40 dilution. Western blot was performed from the immunoprecipitate using ab195289 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

Lane 1 : HeLa whole cell lysate, 10μg (Input).

Lane 2 : ab195289 IP in HeLa whole cell lysate.

Lane 3 : Rabbit IgG,monoclonal [EPR25A]- Isotype Control (ab172730) instead of ab195289 in HeLa whole cell lysate.

Blocking and dilution buffer and concentration : 5% NFDM/TBST.

Exposure time : 3 seconds.

All lanes:

Immunoprecipitation - Anti-USP22 antibody [EPR18945] (ab195289)

Predicted band size: 60 kDa

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• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-USP22 antibody [EPR18945] (AB195289)

Immunohistochemical analysis of paraffin-embedded Rat cerebrum tissue labeling USP22 with ab195289 at 1/1000 followed by a Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) Ready to use. Nuclear staining on rat cerebrum. The section was incubated with ab195289 for 30 mins at room temperature.The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) Ready to use.

Heat mediated antigen retrieval with Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-USP22 antibody [EPR18945] (AB195289)

Immunohistochemical analysis of paraffin-embedded Mouse cerebrum tissue labeling USP22 with ab195289 at 1/1000 followed by a Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) Ready to use. Nuclear staining on mouse cerebrum. The section was incubated with ab195289 for 30 mins at room temperature.The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control : Secondary antibody is Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) Ready to use.

Heat mediated antigen retrieval with Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins

• WB

Lab

Western blot - Anti-USP22 antibody [EPR18945] (AB195289)

Lane 1 : Wild-type HAP1 whole cell lysate (20 μg)
Lane 2 : USP22 knockout HAP1 whole cell lysate (20 μg)
Lane 3 : HEK293 whole cell lysate (20 μg)
Lane 4 : HeLa whole cell lysate (20 μg)

Lanes 1 - 4 : Merged signal (red and green). Green - ab195289 observed at 60 kDa. Red - loading control, ab9484, observed at 37 kDa.

ab195289 was shown to specifically react with USP22 in wild-type HAP1 cells. No band was observed when knockout samples were examined. Wild-type and USP22 knockout samples were subjected to SDS-PAGE. ab195289 and ab9484 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at a 1/2000 dilution and 1/20,000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1/20,000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-USP22 antibody [EPR18945] (ab195289)

Predicted band size: 60 kDa

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• WB

Lab

Western blot - Anti-USP22 antibody [EPR18945] (AB195289)

False colour image of Western blot : Anti-USP22 antibody [EPR18945] staining at 1/2000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab195289 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 [EPR18945] (ab195289) at 1/2000 dilution

Lane 1:

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

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

USP22 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human USP22 knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-usp22-knockout-hela-cell-line-ab264888'>ab264888</a>)

Predicted band size: 60 kDa

Observed band size: 59 kDa

false

• WB

Supplier Data

Western blot - Anti-USP22 antibody [EPR18945] (AB195289)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-USP22 antibody [EPR18945] (ab195289) at 1/2000 dilution

Lane 1:

Human fetal liver lysate at 10 µg

Lane 2:

Human fetal heart lysate at 10 µg

Lane 3:

Human fetal kidney lysate at 10 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/10000 dilution

Predicted band size: 60 kDa

Observed band size: 60 kDa

false

Exposure time: 3min

• WB

Supplier Data

Western blot - Anti-USP22 antibody [EPR18945] (AB195289)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure times : Lane 1, 2, 3, 4, 5 and 6 : 30 seconds; Lane 7 : 15 seconds.

All lanes:

Western blot - Anti-USP22 antibody [EPR18945] (ab195289) at 1/2000 dilution

Lane 1:

HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg

Lane 2:

HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate at 20 µg

Lane 3:

Jurkat (Human T cell leukemia cell line from peripheral blood) whole cell lysate at 20 µg

Lane 4:

HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate at 20 µg

Lane 5:

MCF7 (Human breast adenocarcinoma cell line) whole cell lysate at 20 µg

Lane 6:

Neuro-2a (Mouse neuroblastoma cell line) whole cell lysate at 20 µg

Lane 7:

F9 (Mouse embryonic testicular cancer cell line) whole cell lysate at 20 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 1/100000 dilution

Predicted band size: 60 kDa

Observed band size: 60 kDa

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• WB

Lab

Western blot - Anti-USP22 antibody [EPR18945] (AB195289)

Lane 1 : Wild type HAP1 whole cell lysate (20 μg)
Lane 2 : USP22 knockout HAP1 cell lysate (20 μg)
Lane 3 : HeLa (human cervix adenocarcinoma epithelial cell), whole cell lysate (20 μg)

ab195289 was shown to specifically recognize USP22 when USP22 knockout samples were used. Wild-type and knockout samples were subjected to SDS-PAGE. ab195289 (1/2000) and ab181602 (Rabbit anti GAPDH loading control) were incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1/100,000 dilution.

All lanes:

Western blot - Anti-USP22 antibody [EPR18945] (ab195289)

Predicted band size: 60 kDa

Observed band size: 65 kDa

false

Exposure time: 15s

• WB

Supplier Data

Western blot - Anti-USP22 antibody [EPR18945] (AB195289)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure times : Lane 1, 2, 3 and 4 : 3 minutes; Lane 5, 6, 7 and 8 : 30 seconds.

All lanes:

Western blot - Anti-USP22 antibody [EPR18945] (ab195289) at 1/2000 dilution

Lane 1:

Mouse brain lysate at 10 µg

Lane 2:

Mouse spleen lysate at 10 µg

Lane 3:

Rat brain lysate at 10 µg

Lane 4:

Rat spleen lysate at 10 µg

Lane 5:

C6 (Rat glial tumor cell line) whole cell lysate at 10 µg

Lane 6:

RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate at 10 µg

Lane 7:

PC-12 (Rat adrenal gland pheochromocytoma cell line) whole cell lysate at 10 µg

Lane 8:

NIH/3T3 (Mouse embryonic fibroblast cell line) whole cell lysate at 10 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 1/100000 dilution

Predicted band size: 60 kDa

Observed band size: 60 kDa

false