Anti-Natriuretic peptides A antibody [EPR22089-283]

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Natriuretic peptides A antibody [EPR22089-283] (AB225844)

Immunohistochemical analysis of paraffin-embedded (A) Heart tissue from wild-type C57BL/6JGpt mice and (B) Heart tissue from Nppa knockout mice tissue labeling Natriuretic peptides A with ab225844 at 1/500 dilution, followed by a ready to use LeicaDS9800 (Bond^™ Polymer Refine Detection).

Positive staining on (A) Heart tissue from wild-type C57BL/6JGpt mice and no staining on (B) Heart tissue from Nppa knockout mice.
The section was incubated with ab225844 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 Nppa-KO homozygous mice (Strain ID : T013705).

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-Natriuretic peptides A antibody [EPR22089-283] (AB225844)

Immunohistochemical analysis of paraffin-embedded mouse heart tissue labeling Natriuretic peptides A with ab225844 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Strong cytoplasmic staining mainly in the atria and at lower levels in ventricle of mouse heart (PMID : 2942710; PMID : 25532015; PMID : 1824903). Counterstained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-Fr

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Immunohistochemistry (Frozen sections) - Anti-Natriuretic peptides A antibody [EPR22089-283] (AB225844)

Immunohistochemical analysis of 4% paraformaldehyde-fixed, 0.2% Triton X-100 permeabilized frozen mouse heart tissue labeling Natriuretic peptides A with ab225844 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Cytoplasmic staining in cardiac muscle of mouse atrium is observed.

The nuclear counter stain is DAPI (blue).

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

• IHC-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-Natriuretic peptides A antibody [EPR22089-283] (AB225844)

Immunohistochemical analysis of 4% paraformaldehyde-fixed, 0.2% Triton X-100 permeabilized frozen rat heart tissue labeling Natriuretic peptides A with ab225844 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Cytoplasmic staining in cardiac muscle of rat atrium is observed.

The nuclear counter stain is DAPI (blue).

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Natriuretic peptides A antibody [EPR22089-283] (AB225844)

Immunohistochemical analysis of paraffin-embedded (A) Heart tissue from wild-type C57BL/6JGpt mice and (B) Heart tissue from Nppa knockout mice tissue labeling Natriuretic peptides A with ab225844 at 1/500 dilution followed by a ready to use LeicaDS9800 (Bond^™ Polymer Refine Detection).

Positive staining on (A) Heart tissue from wild-type C57BL/6JGpt mice and no staining on (B) Heart tissue from Nppa knockout mice.
The section was incubated with ab225844 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 Nppa-KO homozygous mice (Strain ID : T013705).

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

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Natriuretic peptides A antibody [EPR22089-283] (AB225844)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized rat cardiomyocyte cells labelling Natriuretic peptides A with ab225844 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^{® :} 488) preadsorbed (ab150081) secondary antibody at 1/1000 dilution.

Confocal image showing cytoplasmic staining in a subset of rat cardiomyocytes (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab9465 Anti-Sarcomeric Alpha Actinin mouse monoclonal antibody was used to counterstain Sarcomeric Alpha Actinin at 1/200 (show in magenta).

The negative controls are as follows :
-ve control 1 : ab225844 at 1/100 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) (ab150120) secondary at 1/1000 dilution.
-ve control 2 : ab9465 at 1/200 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed (ab150081) secondary at 1/1000 dilution.

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Natriuretic peptides A antibody [EPR22089-283] (AB225844)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized mouse cardiomyocyte cells labelling Natriuretic peptides A with ab225844 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^{® :} 488) preadsorbed (ab150081) secondary antibody at 1/1000 dilution.

Confocal image showing cytoplasmic staining in a subset of mouse cardiomyocytes (shown in green). The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab9465 Anti-Sarcomeric Alpha Actinin mouse monoclonal antibody was used to counterstain Sarcomeric Alpha Actinin at 1/200 (show in magenta).

The negative controls are as follows :
-ve control 1 : ab225844 at 1/100 dilution, followed by ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor^® 594) secondary antibody name (ab150120) secondary at 1/1000 dilution.
-ve control 2 : Primary (ab9465) at 1/200 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed (ab150081) secondary at 1/200 dilution.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Natriuretic peptides A antibody [EPR22089-283] (AB225844)

Immunohistochemical analysis of paraffin-embedded rat heart atrium tissue labeling Natriuretic peptides A with ab225844 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining in cardiac muscle of rat atrium (PMID : 2942710; PMID : 25532015; PMID : 1824903). Counterstained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Natriuretic peptides A antibody [EPR22089-283] (AB225844)

Immunohistochemical analysis of paraffin-embedded mouse heart atrium tissue labeling Natriuretic peptides A with ab225844 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining in cardiac muscle of mouse atrium (PMID : 2942710; PMID : 25532015; PMID : 1824903). Counterstained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Heat mediated antigen retrieval was performed with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IP

Supplier Data

Immunoprecipitation - Anti-Natriuretic peptides A antibody [EPR22089-283] (AB225844)

Natriuretic peptides A was immunoprecipitated from 0.35 mg Mouse heart tissue lysate with ab225844 at 1/1000 dilution. Western blot was perfromed from the immunoprecipitate using ab225844 at 1/500 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366) was used for detection at 1/1000 dilution.

Lane 1 : Mouse heart lysate 10 μg (Input).
Lane 2 : ab225844 IP in Mouse heart tissue lysate (+).
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab225844 in Mouse heart lysate (-).

Blocking and dilution buffer and concentration : 5% NFDM/TBST
Exposure time : 10 seconds

All lanes:

Immunoprecipitation - Anti-Natriuretic peptides A antibody [EPR22089-283] (ab225844)

Predicted band size: 16 kDa

false

• WB

Supplier Data

Western blot - Anti-Natriuretic peptides A antibody [EPR22089-283] (AB225844)

Exposure time : Lane 1 : 3 minutes; Lane 2 : 8 seconds.

Blocking/Dilution buffer : 5% NFDM/TBST.

The molecular mass observed is consistent with what has been described in the literature (PMID : 16291870).

All lanes:

Western blot - Anti-Natriuretic peptides A antibody [EPR22089-283] (ab225844) at 1/1000 dilution

Lane 1:

Mouse heart tissue lysate at 10 µg

Lane 2:

Rat heart tissue 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 1/100000 dilution

Predicted band size: 16 kDa

Observed band size: 17 kDa

false

• WB

Lab

Western blot - Anti-Natriuretic peptides A antibody [EPR22089-283] (AB225844)

The tissue samples were kindly provided by GemPharmatech. C57BL/6JGpt wildtype mice and Nppa-KO homozygous mice (Strain ID : T013705).

The identity of the higher MW band at approximately 35 kDa is unknown.

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

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

All lanes:

Western blot - Anti-Natriuretic peptides A antibody [EPR22089-283] (ab225844) at 1/1000 dilution

Lane 1:

Wild-type mouse heart tissue lysate (male)

Lane 2:

Nppa knockout heart tissue lysate (male1)

Lane 3:

Nppa knockout heart tissue lysate (male2)

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: 16 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Natriuretic peptides A antibody [EPR22089-283] (AB225844)

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

ab181602 GAPDH was used as a loading control at 1/1000000 dilution.

Although some references report that ANP is expressed in H9c2 (2-1) cell line (PMID : 31714667, PMID : 30483785), ab225844 is not suitable to detect ANP in this cell line.

All lanes:

Western blot - Anti-Natriuretic peptides A antibody [EPR22089-283] (ab225844) at 1/1000 dilution

Lane 1:

Untreated H9c2 (2-1) (rat cardiac myoblast cell) whole cell lysate at 20 µg

Lane 2:

H9c2 (2-1) (rat cardiac myoblast cell) treated with LPS (2ug/ml, 16h) 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

Predicted band size: 16 kDa

false

Exposure time: 180s