Anti-Claudin 3 antibody [EPR19971]

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Claudin 3 antibody [EPR19971] (AB214487)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed MCF7 (Human breast adenocarcinoma cell line) cells labeling Claudin 3with ab214487 at 1/50 dilution (red) compared withRabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730; black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat Anti-Rabbit IgG (Alexa Fluor^® 488) at 1/2000 dilution was used as the secondary antibody.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Claudin 3 antibody [EPR19971] (AB214487)

Immunohistochemical analysis of paraffin-embedded human prostate tissue labeling Claudin 3 with ab214487 at 1/250 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Membrane staining on the epithelium of human benign prostatic hyperplasia is observed. Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Claudin 3 antibody [EPR19971] (AB214487)

Immunohistochemical analysis of paraffin-embedded human pancreas tissue labeling Claudin 3 with ab214487 at 1/250 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Membrane staining on the pancreatic acinar cells of human pancreas is observed. Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Claudin 3 antibody [EPR19971] (AB214487)

Immunofluorescent analysis of 100% methanol-fixed MCF7 (Human breast adenocarcinoma cell line) cells labeling Claudin 3 with ab214487 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing membrane staining on MCF7 cell line. The nuclear counterstain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) ab195889) at 1/200 dilution (red).

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

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Claudin 3 antibody [EPR19971] (AB214487)

Immunohistochemical analysis of paraffin-embedded human breast cancer tissue labeling Claudin 3 with ab214487 at 1/250 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Membrane staining on the tumor cells of human breast cancer is observed. Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IP

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Immunoprecipitation - Anti-Claudin 3 antibody [EPR19971] (AB214487)

Claudin 3 was immunoprecipitated from 0.35 mg of MCF7 (Human breast adenocarcinoma cell line) whole cell lysate with ab214487 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab214487 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution

Lane 1 : MCF7 whole cell lysate 10μg (Input).

Lane 2 : ab214487 IP in MCF7 whole cell lysate.

Lane 3 : Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) instead of ab214487 in MCF7 whole cell lysate.

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

Exposure time : 3 seconds.

All lanes:

Immunoprecipitation - Anti-Claudin 3 antibody [EPR19971] (ab214487)

Predicted band size: 23 kDa

false

• WB

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Western blot - Anti-Claudin 3 antibody [EPR19971] (AB214487)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1/2 : 1 minute; Lane 3 : 3 seconds; Lane 4 : 3 minutes.

Doublet bands observed correspond to full length and processed forms of the claudin-3 protein. This is consistent with what has been described in the literature (PMID : 26170992).

All lanes:

Western blot - Anti-Claudin 3 antibody [EPR19971] (ab214487) at 1/1000 dilution

Lane 1:

Human breast cancer lysate at 10 µg

Lane 2:

Human pancreas lysate at 10 µg

Lane 3:

Human ovary cancer lysate at 10 µg

Lane 4:

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 1/5000 dilution

Predicted band size: 23 kDa

Observed band size: 20 kDa

false

• WB

Supplier Data

Western blot - Anti-Claudin 3 antibody [EPR19971] (AB214487)

Blocking/Dilution buffer : 5% NFDM/TBST.

Doublet bands observed correspond to full length and processed forms of the claudin-3 protein. This is consistent with what has been described in the literature (PMID : 26170992).

All lanes:

Western blot - Anti-Claudin 3 antibody [EPR19971] (ab214487) at 1/1000 dilution

Lane 1:

SW480 (Human colorectal adenocarcinoma cell line) whole cell lysate at 10 µg

Lane 2:

MCF7 (Human breast adenocarcinoma cell line) whole cell lysate at 10 µg

Lane 3:

Caco-2 (Human colorectal adenocarcinoma 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: 23 kDa

Observed band size: 20 kDa

false

Exposure time: 1s