Anti-ENPP1/PC1 antibody [RM2066]

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-ENPP1/PC1 antibody [RM2066] (AB320007)

Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized HepG2 (human hepatocellular carcinoma epithelial cell) cells labelling ENPP1/PC1 with ab320007 at 1/500 dilution (0.1ug) (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® 647, ab150083) at 1/5000 dilution was used as the secondary antibody.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ENPP1/PC1 antibody [RM2066] (AB320007)

Immunohistochemical analysis of paraffin-embedded Human skeletal muscle tissue labeling ENPP1/PC1 with ab320007 at 1/1000 (0.506 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Low expression : weak staining on human skeletal muscle.
The section was incubated with ab320007 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument
Incubate slides with 3% Hydrogen Peroxide for 10 mins at room temperature after secondary antibody incubation to reduce the background

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

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-ENPP1/PC1 antibody [RM2066] (AB320007)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Tween-20 permeabilized HepG2 (human hepatocellular carcinoma epithelial cell) cells labelling ENPP1/PC1 with ab320007 at 1/1000 (0.506 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 2 ug/ml dilution (Green).

Confocal image showing mainly membranous staining in HepG2 cell line (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue).
Low expression : DLD-1.
Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 2.5 ug/ml dilution (Magenta). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2 ug/ml dilution.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ENPP1/PC1 antibody [RM2066] (AB320007)

Immunohistochemical analysis of paraffin-embedded Human liver tissue labeling ENPP1/PC1 with ab320007 at 1/1000 (0.506 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human liver.
The section was incubated with ab320007 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument
Incubate slides with 3% Hydrogen Peroxide for 10 mins at room temperature after secondary antibody incubation to reduce the background

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-ENPP1/PC1 antibody [RM2066] (AB320007)

ENPP1/PC1 was immunoprecipitated from 0.35 mg Huh7 (human hepatocellular carcinoma epithelial cell) whole cell lysate with ab320007 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab320007 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution.

Lane 1 : Huh7 (human hepatocellular carcinoma epithelial cell) whole cell lysate
Lane 2 : ab320007 IP in Huh7 (human hepatocellular carcinoma epithelial cell) whole cell lysate
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab320007 in Huh7 whole cell lysate

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

All lanes:

Immunoprecipitation - Anti-ENPP1/PC1 antibody [RM2066] (ab320007) at 1/30 dilution

All lanes:

Huh7 (human hepatocellular carcinoma epithelial cell) 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: 76s

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ENPP1/PC1 antibody [RM2066] (AB320007)

Immunohistochemical analysis of paraffin-embedded (A) Liver tissue from wild-type C57BL/6JGpt mice / (B) Liver tissue from ENPP1 knockout mice tissue labeling ENPP1/PC1 with ab320007 at 1/500 dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on (A) Liver tissue from wild-type C57BL/6JGpt mice and no staining on (B) Liver tissue from ENPP1 knockout mice.

The primary antibody was incubated for 30 mins at room temperature.

The immunostaining was performed on a Leica Biosystems BOND^® RX instrument.

The tissue samples were kindly provided by GemPharmatech. C57BL/6JGpt wildtype mice and ENPP1-KO homozygous mice (Strain ID : T013898).

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-ENPP1/PC1 antibody [RM2066] (AB320007)

Immunohistochemical analysis of paraffin-embedded Rat skeletal muscle tissue labeling ENPP1/PC1 with ab320007 at 1/500 (0.506 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Low expression : weak staining on rat skeletal muscle.
The section was incubated with ab320007 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument
Incubate slides with 3% Hydrogen Peroxide for 10 mins at room temperature after secondary antibody incubation to reduce the background

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-ENPP1/PC1 antibody [RM2066] (AB320007)

Immunohistochemical analysis of paraffin-embedded Mouse skeletal muscle tissue labeling ENPP1/PC1 with ab320007 at 1/500 (0.506 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Low expression : weak staining on mouse skeletal muscle.
The section was incubated with ab320007 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument
Incubate slides with 3% Hydrogen Peroxide for 10 mins at room temperature after secondary antibody incubation to reduce the background

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-ENPP1/PC1 antibody [RM2066] (AB320007)

Immunohistochemical analysis of paraffin-embedded Rat liver tissue labeling ENPP1/PC1 with ab320007 at 1/500 (0.506 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on rat liver.
The section was incubated with ab320007 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument
Incubate slides with 3% Hydrogen Peroxide for 10 mins at room temperature after secondary antibody incubation to reduce the background

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-ENPP1/PC1 antibody [RM2066] (AB320007)

Immunohistochemical analysis of paraffin-embedded Mouse liver tissue labeling ENPP1/PC1 with ab320007 at 1/500 (0.506 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on mouse liver.
The section was incubated with ab320007 for 30 mins at room temperature.
The immunostaining was performed on a Leica Biosystems BOND® RX instrument
Incubate slides with 3% Hydrogen Peroxide for 10 mins at room temperature after secondary antibody incubation to reduce the background

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

Supplier Data

Western blot - Anti-ENPP1/PC1 antibody [RM2066] (AB320007)

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

Low expression : DLD-1, Caco-2 (PMID : 31978347), SK-BR-3 (PMID : 23861746).

The identity of the bands higher than 250 kDa and lower than 25 kDa are unknown.

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

Exposure time : Lane 1 : 10 seconds; Lanes 2-5 : 37 seconds.

All lanes:

Western blot - Anti-ENPP1/PC1 antibody [RM2066] (ab320007) at 1/1000 dilution

Lane 1:

Huh7 (human hepatocellular carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

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

Lane 3:

DLD-1 (human colorectal adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

SK-BR-3 (human breast adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 5:

Caco-2 (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='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 130 kDa,36 kDa

false

• WB

Supplier Data

Western blot - Anti-ENPP1/PC1 antibody [RM2066] (AB320007)

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

Low expression : skeletal muscle.

The identity of the bands higher than 250 kDa and lower than 15 kDa are unknown.

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

All lanes:

Western blot - Anti-ENPP1/PC1 antibody [RM2066] (ab320007) at 1/1000 dilution

Lane 1:

Human liver tissue lysate at 20 µg

Lane 2:

Human skeletal muscle tissue lysate at 20 µg

Lane 3:

Human pancreas tissue lysate at 20 µg

Secondary

All lanes:

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

Observed band size: 130 kDa,36 kDa

false

Exposure time: 15s

• WB

Lab

Western blot - Anti-ENPP1/PC1 antibody [RM2066] (AB320007)

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

The tissue samples were kindly provided by GemPharmatech. C57BL/6JGpt wildtype mice and ENPP1-KO homozygous mice (Strain ID : T013898).

This blot was developed using a high-sensitivity ECL substrate, allowing for the detection of proteins in the mid-femtogram range.

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

All lanes:

Western blot - Anti-ENPP1/PC1 antibody [RM2066] (ab320007) at 1/1000 dilution

Lane 1:

Wild-type mouse liver tissue lysate (female) at 20 µg

Lane 2:

ENPP1 knockout mouse liver tissue lysate (female) at 20 µg

Lane 3:

Wild-type mouse liver tissue lysate (male) at 20 µg

Lane 4:

ENPP1 knockout mouse liver tissue lysate (male) at 20 µg

Lane 5:

Wild-type mouse kidney tissue lysate (female) at 20 µg

Lane 6:

ENPP1 knockout mouse kidney tissue lysate (female) at 20 µg

Lane 7:

Wild-type mouse kidney tissue lysate (male) at 20 µg

Lane 8:

ENPP1 knockout mouse kidney tissue lysate (male) 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 1/100000 dilution

Observed band size: 103 kDa,36 kDa

true

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-ENPP1/PC1 antibody [RM2066] (AB320007)

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

All lanes:

Western blot - Anti-ENPP1/PC1 antibody [RM2066] (ab320007) at 1/1000 dilution

Lane 1:

Mouse kidney tissue lysate at 20 µg

Lane 2:

Mouse dorsal ganglion tissue lysate at 20 µg

Lane 3:

Rat dorsal ganglion tissue 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: 130 kDa

false

Exposure time: 81s

• WB

Supplier Data

Western blot - Anti-ENPP1/PC1 antibody [RM2066] (AB320007)

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

Low expression : F9.

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

All lanes:

Western blot - Anti-ENPP1/PC1 antibody [RM2066] (ab320007) at 1/1000 dilution

Lane 1:

NIH/3T3 (mouse embryonic fibroblast) whole cell lysate at 20 µg

Lane 2:

4T1 (mouse mammary gland carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 3:

F9 (mouse embryonal 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: 130 kDa,36 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-ENPP1/PC1 antibody [RM2066] (AB320007)

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

Low expression : skeletal muscle.

The identity of the bands higher than 250 kDa and lower than 50 kDa are unknown.

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

All lanes:

Western blot - Anti-ENPP1/PC1 antibody [RM2066] (ab320007) at 1/1000 dilution

Lane 1:

Mouse liver tissue lysate at 20 µg

Lane 2:

Mouse skeletal muscle tissue lysate at 20 µg

Lane 3:

Rat liver tissue lysate at 20 µg

Lane 4:

Rat skeletal muscle tissue lysate at 20 µg

Lane 5:

Rat kidney tissue 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: 130 kDa,36 kDa

false

Exposure time: 70s

• WB

Supplier Data

Western blot - Anti-ENPP1/PC1 antibody [RM2066] (AB320007)

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

This antibody does not cross-react with mouse ENPP2 and ENPP3.

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

In Western blot, Anti-6X His tag® antibody [EPR20547] - ChIP Grade (ab213204) staining at 1/5000 dilution.

All lanes:

Western blot - Anti-ENPP1/PC1 antibody [RM2066] (ab320007) at 1/1000 dilution

Lane 1:

293T (human embryonic kidney epithelial cell) cells transfected with an empty vector containing a His-tag, whole cell lysate at 10 µg

Lane 2:

293T cells transfected with a mouse ENPP1 expression vector containing a His-tag, whole cell lysate at 10 µg

Lane 3:

293T cells transfected with a mouse ENPP2 expression vector containing a His-tag, whole cell lysate at 10 µg

Lane 4:

293T cells transfected with a mouse ENPP3 expression vector containing a His-tag, 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 1/20000 dilution

Observed band size: 130 kDa,36 kDa

false

Exposure time: 8s