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Phospho-Nucleophosmin (S125) Recombinant Rabbit Monoclonal Antibody [JE51-14]

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Specification

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Catalog# HA722559

Phospho-Nucleophosmin (S125) Recombinant Rabbit Monoclonal Antibody [JE51-14]

Application

  • WB

  • IHC-P

  • IF-Cell

  • FC

Reactivity

  • Human

  • Mouse

  • Rat

Conjugation

  • unconjugated

Overview

Product Name

Phospho-Nucleophosmin (S125) Recombinant Rabbit Monoclonal Antibody [JE51-14]

Antibody Type

Recombinant Rabbit monoclonal Antibody

Immunogen

Synthetic phospho-peptide corresponding to residues surrounding Ser125 of Human Nucleophosmin.

Species Reactivity

Human, Mouse, Rat

Validated Applications

WB, IHC-P, IF-Cell, FC

Target Molecular Weight

Predicted band size: 33 kDa

Positive Control

HeLa cell lysate, NIH/3T3 cell lysate, F9 cell lysate, RAW264.7 cell lysate, L6 cell lysate, PC-12 cell lysate, human kidney tissue, mouse kidney tissue, rat kidney tissue, HeLa, NIH/3T3.

Conjugation

unconjugated

Clone Number

JE51-14

Product Features

Form

Liquid

Concentration

1 mg/mL.(The concentration of this product may be batch-dependent)

Storage Instructions

Shipped at 4℃. Store at +4℃ short term (1-2 weeks). Store at -20℃ long term.

Storage Buffer

1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.

Isotype

IgG

Purification Method

Protein A affinity purified.

Application Dilution

  • WB

  • 1:5,000

  • IHC-P

  • 1:10,000

  • IF-Cell

  • 1:500

  • FC

  • 1:1,000

Target

Function

Nucleophosmin (NPM), also known as nucleolar phosphoprotein B23 or numatrin, is a protein that in humans is encoded by the NPM1 gene. NPM1 is associated with nucleolar ribonucleoprotein structures and binds single-stranded and double-stranded nucleic acids, but it binds preferentially G-quadruplex forming nucleic acids. It is involved in the biogenesis of ribosomes and may assist small basic proteins in their transport to the nucleolus. Its regulation through SUMOylation (by SENP3 and SENP5) is another facet of the protein's regulation and cellular functions. It is located in the nucleolus, but it can be translocated to the nucleoplasm in case of serum starvation or treatment with anticancer drugs. The protein was identified as a phosphoprotein. However, later other post-translational modifications for NPM1 were identified including acetylation and SUMOylation.

Background References

1. Sekhar KR et al. Nucleophosmin Plays a Role in Repairing DNA Damage and Is a Target for Cancer Treatment. Cancer Res. 2023 May

2. Shi Y et al. Nucleophosmin 1: from its pathogenic role to a tantalizing therapeutic target in acute myeloid leukemia. Hematology. 2022 Dec

Subcellular Location

Nucleus, nucleolus, nucleoplasm, Cytoplasm, cytoskeleton, microtubule organizing center, centrosome.

UNIPROT

SwissProt: P06748 Human

SwissProt: Q61937 Mouse

SwissProt: P13084 Rat

Synonyms

B23 antibody

MGC104254 antibody

NO38 antibody

NPM antibody

NPM_HUMAN antibody

NPM1 antibody

Nucleolar phosphoprotein B23 antibody

Nucleolar protein NO38 antibody

Nucleophosmin (nucleolar phosphoprotein B23 numatrin) antibody

Nucleophosmin antibody

Expand

Images

  • Western blot analysis of Phospho-Nucleophosmin (S125) on different lysates with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (<a href="/products/HA722559" style="font-weight: bold;text-decoration: underline;">HA722559</a>) at 1/5,000 dilution.<br /><br />Lane 1: HeLa cell lysate<br />Lane 2: NIH/3T3 cell lysate<br />Lane 3: F9 cell lysate<br />Lane 4: RAW264.7 cell lysate<br />Lane 5: L6 cell lysate<br />Lane 6: PC-12 cell lysate<br /><br />Lysates/proteins at 20 µg/Lane.<br /><br />Predicted band size: 33 kDa<br />Observed band size: 37 kDa<br /><br />Exposure time: 18 seconds; ECL: K1801;<br /><br />4-20% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/HA722559" style="font-weight: bold;text-decoration: underline;">HA722559</a>) at 1/5,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1/50,000 dilution was used for 1 hour at room temperature.

    Western blot analysis of Phospho-Nucleophosmin (S125) on different lysates with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (HA722559) at 1/5,000 dilution.

    Lane 1: HeLa cell lysate
    Lane 2: NIH/3T3 cell lysate
    Lane 3: F9 cell lysate
    Lane 4: RAW264.7 cell lysate
    Lane 5: L6 cell lysate
    Lane 6: PC-12 cell lysate

    Lysates/proteins at 20 µg/Lane.

    Predicted band size: 33 kDa
    Observed band size: 37 kDa

    Exposure time: 18 seconds; ECL: K1801;

    4-20% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA722559) at 1/5,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

  • Immunohistochemical analysis of paraffin-embedded human kidney tissue with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (<a href="/products/HA722559" style="font-weight: bold;text-decoration: underline;">HA722559</a>) at 1/10,000 dilution.<br /><br />The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/HA722559" style="font-weight: bold;text-decoration: underline;">HA722559</a>) at 1/10,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded human kidney tissue with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (HA722559) at 1/10,000 dilution.

    The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA722559) at 1/10,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

  • Immunohistochemical analysis of paraffin-embedded mouse kidney tissue with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (<a href="/products/HA722559" style="font-weight: bold;text-decoration: underline;">HA722559</a>) at 1/10,000 dilution.<br /><br />The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/HA722559" style="font-weight: bold;text-decoration: underline;">HA722559</a>) at 1/10,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded mouse kidney tissue with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (HA722559) at 1/10,000 dilution.

    The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA722559) at 1/10,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

  • Immunohistochemical analysis of paraffin-embedded rat kidney tissue with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (<a href="/products/HA722559" style="font-weight: bold;text-decoration: underline;">HA722559</a>) at 1/10,000 dilution.<br /><br />The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/HA722559" style="font-weight: bold;text-decoration: underline;">HA722559</a>) at 1/10,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded rat kidney tissue with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (HA722559) at 1/10,000 dilution.

    The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA722559) at 1/10,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

  • Immunocytochemistry analysis of HeLa cells labeling Phospho-Nucleophosmin (S125) with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (<a href="/products/HA722559" style="font-weight: bold;text-decoration: underline;">HA722559</a>) at 1/500 dilution.<br /><br />Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (<a href="/products/HA722559" style="font-weight: bold;text-decoration: underline;">HA722559</a>) at 1/500 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor&trade; 488, <a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.<br /><br />Beta tubulin (<a href="/products/M1305-2" style="font-weight: bold;text-decoration: underline;">M1305-2</a>, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor&trade; 594, <a href="/products/HA1126" style="font-weight: bold;text-decoration: underline;">HA1126</a>) was used as the secondary antibody at 1/1,000 dilution.

    Immunocytochemistry analysis of HeLa cells labeling Phospho-Nucleophosmin (S125) with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (HA722559) at 1/500 dilution.

    Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (HA722559) at 1/500 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

    Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

  • Immunocytochemistry analysis of NIH/3T3 cells labeling Phospho-Nucleophosmin (S125) with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (<a href="/products/HA722559" style="font-weight: bold;text-decoration: underline;">HA722559</a>) at 1/500 dilution.<br /><br />Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (<a href="/products/HA722559" style="font-weight: bold;text-decoration: underline;">HA722559</a>) at 1/500 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor&trade; 488, <a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.<br /><br />Beta tubulin (<a href="/products/M1305-2" style="font-weight: bold;text-decoration: underline;">M1305-2</a>, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor&trade; 594, <a href="/products/HA1126" style="font-weight: bold;text-decoration: underline;">HA1126</a>) was used as the secondary antibody at 1/1,000 dilution.

    Immunocytochemistry analysis of NIH/3T3 cells labeling Phospho-Nucleophosmin (S125) with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (HA722559) at 1/500 dilution.

    Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-Nucleophosmin (S125) antibody (HA722559) at 1/500 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

    Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

  • Flow cytometric analysis of HeLa cells labeling Phospho-Nucleophosmin (S125).<br /><br />Cells were fixed and permeabilized. Then stained with the primary antibody (<a href="/products/HA722559" style="font-weight: bold;text-decoration: underline;">HA722559</a>, 1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor&trade; 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (<a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).

    Flow cytometric analysis of HeLa cells labeling Phospho-Nucleophosmin (S125).

    Cells were fixed and permeabilized. Then stained with the primary antibody (HA722559, 1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).

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