Anti-GPA33 抗体 [EPR4240]

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-GPA33 antibody [EPR4240] (AB108938)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized LoVo (human colorectal adenocarcinoma epithelial cell) cells labelling GPA33 with ab108938 at 1/50 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed (ab150081) antibody at 1/1000 dilution (Green).

Confocal image showing membrane and weak cytoplasmic staining in LoVo cell line (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue).

Low expression : Caco2.

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

Anti-alpha Tubulin mouse monoclonal antibody (ab7291) was used to counterstain tubulin at 1/1000 dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed (ab150081) at 1/1000 dilution.

• WB

Unknown

Western blot - Anti-GPA33 antibody [EPR4240] (AB108938)

All lanes:

Western blot - Anti-GPA33 antibody [EPR4240] (ab108938) at 1/1000 dilution

Lane 1:

HT-29 lysate at 10 µg

Lane 2:

Human small intestine lysate at 10 µg

Lane 3:

Human fetal colon lysates at 10 µg

Secondary

All lanes:

HRP-labelled goat anti-rabbit at 1/2000 dilution

Predicted band size: 36 kDa

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• WB

Lab

Western blot - Anti-GPA33 antibody [EPR4240] (AB108938)

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

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

Low expression : Caco-2.

The identity of the lower MW bands at approximately 25 kDa (in lane 1 and 3) are unknown.

All lanes:

Western blot - Anti-GPA33 antibody [EPR4240] (ab108938) at 1/1000 dilution

Lane 1:

Caco-2 (human colorectal adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

HCT 116 (human colorectal carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 3:

LoVo (human colorectal adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

THP-1 (human monocytic leukemia monocyte) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 35 kDa

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Exposure time: 92s

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GPA33 antibody [EPR4240] (AB108938)

Unpurified ab108938 showing positive staining in Ovarian carcinoma tissue.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GPA33 antibody [EPR4240] (AB108938)

Unpurified ab108938 at 1/250 dilution staining GPA33 in paraffin-embedded Human colonic adenocarcinoma tissue by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections).

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GPA33 antibody [EPR4240] (AB108938)

Unpurified ab108938 showing negative staining in Breast carcinoma tissue.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GPA33 antibody [EPR4240] (AB108938)

Unpurified ab108938 showing negative staining in Normal tonsil tissue.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GPA33 antibody [EPR4240] (AB108938)

Unpurified ab108938 showing negative staining in Thyroid gland carcinoma tissue.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GPA33 antibody [EPR4240] (AB108938)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human colon cancer tissue sections labeling GPA33 with Purified ab108938 at 1 : 400 dilution (0.25 μg/ml). Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control : PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GPA33 antibody [EPR4240] (AB108938)

Unpurified ab108938 showing negative staining in Normal liver tissue.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-GPA33 antibody [EPR4240] (AB108938)

Unpurified ab108938 at 1/250 dilution staining GPA33 in paraffin-embedded Human colon tissue by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections).

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• IP

Lab

Immunoprecipitation - Anti-GPA33 antibody [EPR4240] (AB108938)

Purified ab108938 at 1 : 20 dilution (0.5ug) immunoprecipitating GPA33 in HT-29 whole cell lysate.

Lane 1 (input) : HT-29 (Human colorectal adenocarcinoma epithelial cell) whole cell lysate (10μg)

Lane 2 (+) : ab108938 + HT-29 whole cell lysate.

Lane 3 (-) : Rabbit monoclonal IgG (ab172730) instead of ab108938 in HT-29 whole cell lysate.

VeriBlot for IP Detection Reagent (HRP) (ab131366) (1/1000) was used for Western blotting.

Blocking Buffer and concentration : 5% NFDM/TBST.

Diluting buffer and concentration : 5% NFDM/TBST.

Observed band size : 50 kDa

All lanes:

Immunoprecipitation - Anti-GPA33 antibody [EPR4240] (ab108938)

Predicted band size: 36 kDa

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• WB

Unknown

Western blot - Anti-GPA33 antibody [EPR4240] (AB108938)

Blocking/Diluting buffer : 5% NFDM/TBST

All lanes:

Western blot - Anti-GPA33 antibody [EPR4240] (ab108938) at 1/1000 dilution

Lane 1:

Human colon lysates at 20 µg

Lane 2:

HT-29 (Human colorectal adenocarcinoma epithelial cell) whole cell lysates at 20 µg

Lane 3:

MOLT-4 (Human lymphoblastic leukemia T lymphoblast) whole cell lysates at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Predicted band size: 36 kDa

Observed band size: 50 kDa

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• WB

CiteAb

Western blot - Anti-GPA33 antibody [EPR4240] (AB108938)

GPA33 western blot using anti-GPA33 antibody [EPR4240] ab108938. Publication image and figure legend from Gotanda, K., Hirota, T., et al., 2016, Sci Rep, PubMed 27571936.

ab108938 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab108938 please see the product overview.

Immunoprecipitation experiments confirming the identification of intestine-derived exosomes.(A) GPA33 proteins in total exosomes and intestine-derived exosomes were detected by Western blot. CD9 is a marker of exosomes. (B) Tissue-specific miRNA levels in intestine-derived exosomes. MiR-192 and miR-215 are intestine-specific miRNAs and miR-122 is a liver-specific miRNA. Relative miRNA levels were determined by qRT-PCR. Each column represents the mean ± S.D. (n = 3). *p < 0.05, significantly different from the input. Input : total exosome samples, IP : Immunoprecipitated samples.

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