Anti-Lewis a,b carbohydrate antibody [HEA164]

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lewis a,b carbohydrate antibody [HEA164] (AB322960)

Immunohistochemical analysis of paraffin-embedded Human liver tissue labeling Lewis a,b carbohydrate with ab322960 at 1/10000 dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Low expression : no staining on human liver.
The section was incubated with ab322960 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 a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Lewis a,b carbohydrate antibody [HEA164] (AB322960)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HepG2 (human hepatocellular carcinoma epithelial cell, Left) / BxPC-3 (human pancreas adenocarcinoma epithelial cell, Right) cells labeling Lewis a,b carbohydrate with ab322960 at 1/100 dilution (1ug) (Red) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (Blue).

Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/5000 dilution was used as the secondary antibody.

Negative control : HepG2

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lewis a,b carbohydrate antibody [HEA164] (AB322960)

Immunohistochemical analysis of paraffin-embedded Human pancreas cancer tissue labeling Lewis a,b carbohydrate with ab322960 at 1/10000 dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human pancreas cancer.
The section was incubated with ab322960 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 a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Lewis a,b carbohydrate antibody [HEA164] (AB322960)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized BxPC-3 (human pancreas adenocarcinoma epithelial cell) cells labelling Lewis a,b carbohydrate with ab322960 at 1/2000 dilution, followed by ab150113 Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) antibody at 1/1000 dilution (Green).

Confocal image showing cytoplasmic and membranous staining in BxPC-3 cell line (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue).
Negative control : HepG2.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab206369 Anti-beta Tubulin rabbit monoclonal antibody (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150113 Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) at 1/1000 dilution.

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lewis a,b carbohydrate antibody [HEA164] (AB322960)

Immunohistochemical analysis of paraffin-embedded Human colon cancer tissue labeling Lewis a,b carbohydrate with ab322960 at 1/10000 dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human colon cancer.
The section was incubated with ab322960 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 a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Lewis a,b carbohydrate antibody [HEA164] (AB322960)

Immunohistochemical analysis of paraffin-embedded Human colon tissue labeling Lewis a,b carbohydrate with ab322960 at 1/10000 dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human colon.
The section was incubated with ab322960 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 a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Heat mediated antigen retrieval was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• WB

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Western blot - Anti-Lewis a,b carbohydrate antibody [HEA164] (AB322960)

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

Negative control : HepG2
Low expression tissue : human liver

The identity of the lower MW band at approximately 150 kDa (in lane 3) is unknown.

Exposure time : Lane 1 : 8 seconds; Lane 2-4 : 59 seconds

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

All lanes:

Western blot - Anti-Lewis a,b carbohydrate antibody [HEA164] (ab322960) at 1/1000 dilution

Lane 1:

BxPC-3 (human pancreas adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 3:

Human pancreas cancer tissue lysate at 20 µg

Lane 4:

Human liver tissue lysate at 20 µg

Secondary

All lanes:

Peroxidase-Conjugated Goat anti-Mouse IgG (H+L) at 1/10000 dilution

Observed band size: 250 kDa,36 kDa

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