Anti-Tspan-8 antibody [EPR28888-508]

• Flow Cyt

Lab

Flow Cytometry - Anti-Tspan-8 antibody [EPR28888-508] (AB326570)

Flow cytometric analysis of LS513 (human Large intestine Cecum epithelial cell, Right) Daudi (human Burkitt's lymphoma lymphoblast, Left) cells labelling Tspan-8 with ab326570 at 1/50 dilution (1ug) / Red compared with a Rabbit monoclonal IgG (ab172730) / Black isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody).

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

Gated on viable cells. Low expression : Daudi.

• mIHC

Lab

Multiplex immunohistochemistry - Anti-Tspan-8 antibody [EPR28888-508] (AB326570)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded human small intestine tissue staining SMOC2 with ab323479 at a 1/100 dilution (4.86 μg/ml), ab326570 anti-Tspan-8 at a 1/100 dilution (4.84 μg/ml) and ab272692 anti-MUC2 at a 1/2000 dilution (0.253 μg/ml).

Panel A : merged staining of anti-SMOC2 (green; Opal™520), anti-Tspan-8 (magenta; Opal™690) and anti-MUC2 (gray; Opal™570) on human small intestine.

Panel B : anti-SMOC2 staining stem cells in human small intestine.

Panel C : anti-Tspan-8 staining epithelium in human small intestine.

Panel D : anti-MUC2 staining goblet cells in human small intestine.

Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab323479, ab326570 and ab272692 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND^® RX instrument with an Opal™ 4-color kit using Opal Polymer HRP Ms + Rb secondary reagent. Image acquisition was performed with a confocal microscope (Leica-Microsystems, STELLARIS).

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

• mIHC

Lab

Multiplex immunohistochemistry - Anti-Tspan-8 antibody [EPR28888-508] (AB326570)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded human colon tissue staining Fibrillin 1 with ab314547 at a 1/100 dilution (5.02 μg/ml), ab326570 anti-Tspan-8 used at a 1/100 dilution (4.84 μg/ml) and ab272692 anti-MUC2 used at a 1/2000 dilution (0.253 μg/ml).

Panel A : merged staining of anti-Fibrillin 1 (green; Opal™520), anti-Tspan-8 (magenta; Opal™690) and anti-MUC2 (gray; Opal™570) on human colon.

Panel B : anti-Fibrillin 1 staining extracellular microfibrils in human colon.

Panel C : anti-Tspan-8 staining epithelium in human colon.

Panel D : anti-MUC2 staining goblet cells in human colon.

Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab314547, ab326570 and ab272692 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND^® RX instrument with an Opal™ 4-color kit using Opal Polymer HRP Ms + Rb secondary reagent. Image acquisition was performed with Leica SP8 confocal microscope.

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

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Tspan-8 antibody [EPR28888-508] (AB326570)

Immunohistochemical analysis of paraffin-embedded Human epityphlon tissue labeling Tspan-8 with ab326570 at 1/100 (4.84 µg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human epityphlon. The primary antibody was incubated 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

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Tspan-8 antibody [EPR28888-508] (AB326570)

Immunohistochemical analysis of paraffin-embedded Human colon tissue labeling Tspan-8 with ab326570 at 1/100 (4.84 µg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human colon. The primary antibody was incubated 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

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Tspan-8 antibody [EPR28888-508] (AB326570)

Immunohistochemical analysis of paraffin-embedded Human liver carcinoma tissue labeling Tspan-8 with ab326570 at 1/100 (4.84 µg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human liver carcinoma. The primary antibody was incubated 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

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Tspan-8 antibody [EPR28888-508] (AB326570)

Immunohistochemical analysis of paraffin-embedded Human pancreatic carcinoma tissue labeling Tspan-8 with ab326570 at 1/100 (4.84 µg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human pancreatic carcinoma. The primary antibody was incubated 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

• mIHC

Lab

Multiplex immunohistochemistry - Anti-Tspan-8 antibody [EPR28888-508] (AB326570)

Multiplex immunohistochemistry analysis of formalin/PFA-fixed paraffin-embedded human colon tissue staining SMOC2 with ab323479 at a 1/100 dilution (4.86 μg/ml), ab326570 anti-Tspan-8 at a 1/100 dilution (4.84 μg/ml) and ab272692 anti-MUC2 at a 1/2000 dilution (0.253 μg/ml).

Panel A : merged staining of anti-SMOC2 (green; Opal™520), anti-Tspan-8 (magenta; Opal™690) and anti-MUC2 (gray; Opal™570) on human colon.

Panel B : anti-SMOC2 staining stem cells in human colon.

Panel C : anti-Tspan-8 staining epithelium in human colon.

Panel D : anti-MUC2 staining goblet cells in human colon.

Nuclear DNA was labeled with DAPI (shown in blue).

The section was incubated in three rounds of staining : in the order of ab323479, ab326570 and ab272692 for 30 mins at room temperature. Each round was followed by a separate fluorescent tyramide signal amplification system.

The immunostaining was performed on a Leica Biosystems BOND^® RX instrument with an Opal™ 4-color kit using Opal Polymer HRP Ms + Rb secondary reagent. Image acquisition was performed with a Leica STELLARIS confocal microscope.

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

• I-ELISA

Lab

Indirect ELISA - Anti-Tspan-8 antibody [EPR28888-508] (AB326570)

Indirect ELISA analysis of ab326570 at 1000-0 ng/ml. The Secondary antibody used was Alkaline Phosphatase-conjugated AffiniPure Goat Anti-Rabbit IgG (H+L) at 1 : 2500 dilution dilution.

Antigen : Human Tspan-8 (extracellular domain fragment);Human Tspan-1 (extracellular domain fragment);Human Tspan-9 (extracellular domain fragment).

Antigen concentration : 1000 ng/ml

This antibody does not cross-react with recombinant human Tspan-1 or Tspan-9 by ELISA.

• WB

Lab

Western blot - Anti-Tspan-8 antibody [EPR28888-508] (AB326570)

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

Negative control : human skeletal muscle, human spleen.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) (1 : 200000) (36KDa).

All lanes:

Western blot - Anti-Tspan-8 antibody [EPR28888-508] (ab326570) at 1/1000 dilution

Lane 1:

Human stomach tissue lysate at 30 µg

Lane 2:

Human skeletal muscle tissue lysate at 30 µg

Lane 3:

Human spleen tissue lysate at 30 µg

Lane 4:

Human colon tissue lysate at 30 µg

Secondary

All lanes:

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

Observed band size: 26 kDa,36 kDa

false

Exposure time: 180s

• WB

Lab

Western blot - Anti-Tspan-8 antibody [EPR28888-508] (AB326570)

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

Negative control : Daudi, MCF7.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) (1 : 200000) (36KDa).

All lanes:

Western blot - Anti-Tspan-8 antibody [EPR28888-508] (ab326570) at 1/1000 dilution

Lane 1:

AsPC-1 (human Pancreas epithelial cell) whole cell lysate at 20 µg

Lane 2:

LS513 (human Large intestine, Cecum epithelial cell) whole cell lysate at 20 µg

Lane 3:

Daudi (human burkitt's lymphoma lymphoblast) whole cell lysate at 20 µg

Lane 4:

MCF7 (human breast adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 5:

SW1463 (human Large intestine, Rectum epithelial cell) whole cell lysate at 20 µg

Lane 6:

HT115 (human colon epithelial cell) whole cell lysate at 20 µg

Lane 7:

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

Lane 8:

SW1116 (human colorectal adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Secondary

All lanes:

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

Observed band size: 26 kDa,36 kDa

false

Exposure time: 180s

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Tspan-8 antibody [EPR28888-508] (AB326570)

Immunohistochemical analysis of paraffin-embedded Human spleen tissue labeling Tspan-8 with ab326570 at 1/100 (4.84 µg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : no staining on human spleen. The primary antibody was incubated 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

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Tspan-8 antibody [EPR28888-508] (AB326570)

Immunohistochemical analysis of paraffin-embedded Human skeletal muscle tissue labeling Tspan-8 with ab326570 at 1/100 (4.84 µg/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : no staining on human skeletal muscle. The primary antibody was incubated 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