Anti-HHLA2 antibody [EPR26395-88]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HHLA2 antibody [EPR26395-88] (AB315994)

Immunohistochemical analysis of paraffin-embedded (A) HEK-293T (human embryonic kidney epithelial cell) transfected with a human HHLA2 expression vector containing a Myc-His tag. (B) HEK-293T transfected with empty vector containing a Myc-His tag. Tissue labeling HHLA2 with ab315994 at 1/500 (0.982 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on HEK-293T transfected with a Myc-His-tagged human HHLA2 construct (image A). No staining on HEK-293T transfected with empty plasmid (image B). The section was incubated with ab315994 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-HHLA2 antibody [EPR26395-88] (AB315994)

Immunohistochemical analysis of paraffin-embedded Human spleen tissue labeling HHLA2 with ab315994 at 1/500 (0.982 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Negative control : no staining on human spleen (PMID : 32071413). The section was incubated with ab315994 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-HHLA2 antibody [EPR26395-88] (AB315994)

Immunohistochemical analysis of paraffin-embedded Human colon carcinoma tissue labeling HHLA2 with ab315994 at 1/500 (0.982 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human colon carcinoma (PMID : 25549724). The section was incubated with ab315994 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-HHLA2 antibody [EPR26395-88] (AB315994)

Immunohistochemical analysis of paraffin-embedded (A) A-204 (human muscle rhabdomyosarcoma cell) cell pellet (B) HeLa (human cervical adenocarcinoma epithelial cell) cell pellet tissue labeling HHLA2 with ab315994 at 1/500 (0.982 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on A-204 (image A). No staining on Hela (image B). The section was incubated with ab315994 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-HHLA2 antibody [EPR26395-88] (AB315994)

Immunohistochemical analysis of paraffin-embedded Human colon tissue labeling HHLA2 with ab315994 at 1/500 (0.982 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human colon (PMID : 25549724). The section was incubated with ab315994 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.

• IP

Supplier Data

Immunoprecipitation - Anti-HHLA2 antibody [EPR26395-88] (AB315994)

HHLA2 was immunoprecipitated from 0.35 mg A-204 (human muscle rhabdomyosarcoma cell) whole cell lysate with ab315994 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab315994 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

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

All lanes:

Immunoprecipitation - Anti-HHLA2 antibody [EPR26395-88] (ab315994) at 1/1000 dilution

Lane 1:

A-204 (human muscle rhabdomyosarcoma cell) whole cell lysate

Lane 2:

ab315994 at 1/30 IP in A-204 (human muscle rhabdomyosarcoma cell) whole cell lysate

Lane 3:

Rabbit monoclonal IgG (<a href='/en-us/products/primary-antibodies/rabbit-igg-monoclonal-epr25a-isotype-control-ab172730'>ab172730</a>) instead of ab315994 in A-204 whole cell lysate

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-HHLA2 antibody [EPR26395-88] (AB315994)

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

Negative control : HeLa, Jurkat (PMID : 25549724), HepG2.

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

Exposure time : Lane 1 : 37 seconds; Lanes 2-7 : 10 seconds.

All lanes:

Western blot - Anti-HHLA2 antibody [EPR26395-88] (ab315994) at 1/1000 dilution

Lane 1:

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

Lane 2:

HCC827 (human lung adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 3:

HDLM-2 (human hodgkin lymphoma cell) whole cell lysate at 20 µg

Lane 4:

A-204 (human muscle rhabdomyosarcoma cell) whole cell lysate at 20 µg

Lane 5:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 6:

Jurkat (human t cell leukemia t lymphocyte from peripheral blood) whole cell lysate at 20 µg

Lane 7:

HepG2 (human hepatocellar carcinoma epithelial cell) whole cell lysate at 20 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 1/20000 dilution

Observed band size: 70-80 kDa,36 kDa

false