Anti-CD137 antibody [EPR23218-111]

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-CD137 antibody [EPR23218-111] (AB243648)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HEK-293T cells labelling CD137 with ab243648 at 1/100 dilution, followed by ab150080 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 594) antibody at 1/1000 dilution (Red). Confocal image showing cytoplasmic and membranous staining in HEK-293T cells transfected with a GFP-tagged CD137 expression vector - the GPF tag is shown in Green. The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150080 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 594) at 1/1000 dilution.

• Flow Cyt

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Flow Cytometry - Anti-CD137 antibody [EPR23218-111] (AB243648)

Flow cytometric analysis of Mouse splenocytes (treated with 2.5ug/ml concanavalin A for 72h) cells labelling CD137 with ab243648 at 1/500 (Right) compared with a Rabbit monoclonal IgG (ab172730) isotype control (Left).

Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.

Cells were stained with rabbit IgG (Left) or ab243648 (Right), then stained with anti-CD25 conjugated to BV421.

Gated on viable cells.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-CD137 antibody [EPR23218-111] (AB243648)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized Mouse splenocytes cells labelling CD137 with ab243648 at 1/100 dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic and membranous staining in mouse splenocytes treated with Concanavalin A (2.5 ug/ml) for 72 h. ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution.

• IP

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Immunoprecipitation - Anti-CD137 antibody [EPR23218-111] (AB243648)

CD137 was immunoprecipitated from 0.35 mg mouse splenocyte (treated with 2.5 μg/ml Concanavalin A for 72 hours) whole cell lysate with ab243648 at 1/30 dilution. Western blot was performed on the immunoprecipitate using ab243648 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : Mouse splenocyte (treated with 2.5 μg/ml Concanavalin A for 72 hours) whole cell lysate 10 μg

Lane 2 : ab243648 IP in Mouse splenocyte (treated as above) whole cell lysate

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab243648 in Mouse splenocyte (treated as above) whole cell lysate

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

Exposure time : 76 seconds.

CD137 is a type I transmembrane glycoprotein and has both monomeric and dimeric forms on the surface of activated T cells (PMID : 30355497).

All lanes:

Immunoprecipitation - Anti-CD137 antibody [EPR23218-111] (ab243648)

Predicted band size: 28 kDa,63 kDa

Observed band size: 21-24 kDa,36-40 kDa

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• WB

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Western blot - Anti-CD137 antibody [EPR23218-111] (AB243648)

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

CD137 is a type I transmembrane glycoprotein and has both monomeric and dimeric forms on the surface of activated T cells (PMID : 30355497).

Exposure time : 3 minutes.

All lanes:

Western blot - Anti-CD137 antibody [EPR23218-111] (ab243648) at 1/1000 dilution

Lane 1:

Untreated mouse splenocytes, whole cell lysate at 20 µg

Lane 2:

Mouse splenocytes (treated with 2.5 μg/ml Concanavalin A for 72 hours), 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 1/50000 dilution

Predicted band size: 28 kDa

Observed band size: 21-24 kDa,36-40 kDa

false

• WB

Unknown

Western blot - Anti-CD137 antibody [EPR23218-111] (AB243648)

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

Exposure time : 15 seconds.

All lanes:

Western blot - Anti-CD137 antibody [EPR23218-111] (ab243648) at 1/1000 dilution

Lane 1:

His-tagged mouse CD137 recombinant protein(aa24-187), 10 ng

Lane 2:

His-tagged human CD137 recombinant protein(aa24-186), 10 ng

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: 28 kDa

false