Anti-Claudin 1 antibody [EPRR18871]

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Claudin 1 antibody [EPRR18871] (AB211737)

Immunohistochemical analysis of paraffin-embedded human colon cancer tissue labeling Claudin 1 with ab211737 at 1/250 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Membrane staining on human colon cancer (PMID : 15475928) 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

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Claudin 1 antibody [EPRR18871] (AB211737)

Immunohistochemical analysis of paraffin-embedded human cervix tissue labeling Claudin 1 with ab211737 at 1/250 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Membrane staining on human cervix 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 1 antibody [EPRR18871] (AB211737)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HepG2 (Human liver hepatocellular carcinoma cell line) cells labeling Claudin 1 with ab211737 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing membranous staining on HepG2 cell line.

The nuclear counterstain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) (ab150120) secondary antibody at 1/1000 dilution (red).

The negative controls are as follows : -

-ve control 1 : ab211737 at 1/1000 dilution followed by Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) (ab150120) secondary antibody at 1/1000 dilution.

-ve control 2 : Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Claudin 1 antibody [EPRR18871] (AB211737)

Immunohistochemical analysis of paraffin-embedded human lung adenocarcinoma tissue labeling Claudin 1 with ab211737 at 1/250 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Negative control : Negative staining on human lung adenocarcinoma, which is consistent with the literature (PMID : 17585317).

Counter stained with Hematoxylin.

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

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Claudin 1 antibody [EPRR18871] (AB211737)

Immunohistochemical analysis of paraffin-embedded human skin tissue labeling Claudin 1 with ab211737 at 1/250 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Membrane staining on human skin squamous cells 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.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Claudin 1 antibody [EPRR18871] (AB211737)

Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed HepG2 (Human liver hepatocellular carcinoma cell line) cells labeling Claudin 1with ab211737 at 1/60 dilution (red) compared with a rabbit monoclonal IgG 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.

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Claudin 1 antibody [EPRR18871] (AB211737)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized A431 (Human epidermoid carcinoma cell line) cells labeling Claudin 1 with ab211737 at 1/1000 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 A431 cell line.The nuclear counterstain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) (ab150120) secondary antibody at 1/1000 dilution (red).

The negative controls are as follows : -

-ve control 1 : ab211737 at 1/1000 dilution followed by Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) (ab150120) secondary antibody at 1/1000 dilution.

-ve control 2 : Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Claudin 1 antibody [EPRR18871] (AB211737)

Immunohistochemical analysis of paraffin-embedded human lung squamous cell cancer tissue labeling Claudin 1 with ab211737 at 1/250 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution. Membrane staining on human lung squamous cell 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 1 antibody [EPRR18871] (AB211737)

Claudin 1 was immunoprecipitated from 0.35 mg of HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate with ab211737 at 1/40 dilution.

Western blot was performed from the immunoprecipitate using ab211737 at 1/2000 dilution.

VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10,000 dilution

Lane 1 : HepG2 whole cell lysate 10μg (Input).

Lane 2 : ab211737 IP in HepG2 whole cell lysate.

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab211737 in HepG2 whole cell lysate.

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

Exposure time : 3 seconds.

All lanes:

Immunoprecipitation - Anti-Claudin 1 antibody [EPRR18871] (ab211737)

Predicted band size: 22 kDa

false

• WB

Supplier Data

Western blot - Anti-Claudin 1 antibody [EPRR18871] (AB211737)

Blocking/Dilution buffer : 5% NFDM/TBST.

The expression profile/ molecular weight observed is consistent with what has been described in the literature PMID : 26601075.

All lanes:

Western blot - Anti-Claudin 1 antibody [EPRR18871] (ab211737) at 1/2000 dilution

Lane 1:

Human fetal liver lysate at 10 µg

Lane 2:

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: 22 kDa

Observed band size: 18 kDa

false

Exposure time: 3min

• WB

Lab

Western blot - Anti-Claudin 1 antibody [EPRR18871] (AB211737)

Lanes 1 - 4 : Merged signal (red and green). Green - ab211737 observed at 18 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab211737 was shown to specifically react with in wild-type A431 WT cells as signal was lost in A431 CLDN1 knockout cells. Wild-type and A431 CLDN1 knockout samples were subjected to SDS-PAGE. The membrane was blocked with 3pc Milk. ab211737 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1/2000 dilution and 1/20000 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 1/20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-Claudin 1 antibody [EPRR18871] (ab211737) at 1/2000 dilution

Lane 1:

Wild-type A-431 (Human epidermoid carcinoma cell line) whole cell lysate at 20 µg

Lane 2:

CLDN1 knockout A-431 (Human epidermoid carcinoma cell line) whole cell lysate at 20 µg

Lane 2:

Western blot - Human CLDN1 knockout A-431 cell line (<a href='/en-us/products/cell-lines/human-cldn1-knockout-a-431-cell-line-ab261889'>ab261889</a>)

Lane 3:

A549 (Human lung carcinoma cell line) whole cell lysate at 20 µg

Lane 4:

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

Predicted band size: 22 kDa

false

• WB

Supplier Data

Western blot - Anti-Claudin 1 antibody [EPRR18871] (AB211737)

Blocking/Dilution buffer : 5% NFDM/TBST.

Human Claudin 1 full length recombinant protein contains aa1-211 with His-Tag® and GST-tag. This recombinant was made in house.

All lanes:

Western blot - Anti-Claudin 1 antibody [EPRR18871] (ab211737) at 1/5000 dilution

All lanes:

Human Claudin 1 full length recombinant protein at 0.01 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: 22 kDa

Observed band size: 45 kDa

false

Exposure time: 1s

• WB

Supplier Data

Western blot - Anti-Claudin 1 antibody [EPRR18871] (AB211737)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure time : Lane 1/2 : 15 seconds; Lane 3 : 5 seconds.

The expression profile/ molecular weight observed is consistent with what has been described in the literature PMID : 26601075.

All lanes:

Western blot - Anti-Claudin 1 antibody [EPRR18871] (ab211737) at 1/2000 dilution

Lane 1:

HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate at 10 µg

Lane 2:

A431 (Human epidermoid carcinoma cell line) whole cell lysate at 10 µg

Lane 3:

HepG2 (Human liver hepatocellular carcinoma 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: 22 kDa

Observed band size: 18 kDa

false