Overview

  • Product name
    Anti-Nucleophosmin antibody
    See all Nucleophosmin primary antibodies
  • Description
    Rabbit polyclonal to Nucleophosmin
  • Host species
    Rabbit
  • Tested applications
    Suitable for: IHC-P, ICC/IF, WBmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    This product was produced with the following immunogens:
    Synthetic peptide corresponding to Human Nucleophosmin aa 23-38.

    Synthetic peptide corresponding to Human Nucleophosmin aa 226-240.

  • Positive control
    • HeLa or mouse fibroblast extract.

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C. Avoid freeze / thaw cycle.
  • Storage buffer
    Preservative: 0.05% Sodium azide
    Constituents: 99% PBS, 0.1% BSA
  • Concentration information loading...
  • Purity
    Immunogen affinity purified
  • Clonality
    Polyclonal
  • Isotype
    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab15440 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
IHC-P 1/200. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
ICC/IF 1/100. Used at a dilution of 1/100 (see Abreview for further information).
WB 1/500 - 1/1000. Detects a band of approximately 38 kDa (predicted molecular weight: 33 kDa).

Target

  • Function
    Involved in diverse cellular processes such as ribosome biogenesis, centrosome duplication, protein chaperoning, histone assembly, cell proliferation, and regulation of tumor suppressors p53/TP53 and ARF. Binds ribosome presumably to drive ribosome nuclear export. Associated with nucleolar ribonucleoprotein structures and bind single-stranded nucleic acids. Acts as a chaperonin for the core histones H3, H2B and H4. Stimulates APEX1 endonuclease activity on apurinic/apyrimidinic (AP) double-stranded DNA but inhibits APEX1 endonuclease activity on AP single-stranded RNA. May exert a control of APEX1 endonuclease activity within nucleoli devoted to repair AP on rDNA and the removal of oxidized rRNA molecules. In concert with BRCA2, regulates centrosome duplication. Regulates centriole duplication: phosphorylation by PLK2 is able to trigger centriole replication. Negatively regulates the activation of EIF2AK2/PKR and suppresses apoptosis through inhibition of EIF2AK2/PKR autophosphorylation. Antagonizes the inhibitory effect of ATF5 on cell proliferation and relieves ATF5-induced G2/M blockade (PubMed:22528486).
  • Involvement in disease
    A chromosomal aberration involving NPM1 is found in a form of non-Hodgkin lymphoma. Translocation t(2;5)(p23;q35) with ALK. The resulting chimeric NPM1-ALK protein homodimerize and the kinase becomes constitutively activated.
    A chromosomal aberration involving NPM1 is found in a form of acute promyelocytic leukemia. Translocation t(5;17)(q32;q11) with RARA.
    A chromosomal aberration involving NPM1 is a cause of myelodysplastic syndrome (MDS). Translocation t(3;5)(q25.1;q34) with MLF1.
    Defects in NPM1 are associated with acute myelogenous leukemia (AML). Mutations in exon 12 affecting the C-terminus of the protein are associated with an aberrant cytoplasmic location.
  • Sequence similarities
    Belongs to the nucleoplasmin family.
  • Post-translational
    modifications
    Acetylated at C-terminal lysine residues, thereby increasing affinity to histones.
    ADP-ribosylated.
    Phosphorylated at Ser-4 by PLK1 and PLK2. Phosphorylation at Ser-4 by PLK2 in S phase is required for centriole duplication and is sufficient to trigger centriole replication. Phosphorylation at Ser-4 by PLK1 takes place during mitosis. Phosphorylated by CDK2 at Ser-125 and Thr-199. Phosphorylation at Thr-199 may trigger initiation of centrosome duplication. Phosphorylated by CDK1 at Thr-199, Thr-219, Thr-234 and Thr-237 during cell mitosis. When these four sites are phosphorated, RNA-binding activity seem to be abolished. May be phosphorylated at Ser-70 by NEK2. The Thr-199 phosphorylated form has higher affinity for ROCK2. CDK6 triggers Thr-199 phosphorylation when complexed to Kaposi's sarcoma herpesvirus (KSHV) V-cyclin, leading to viral reactivation by reducing viral LANA levels.
    Sumoylated by ARF.
  • Cellular localization
    Nucleus, nucleolus. Nucleus, nucleoplasm. Cytoplasm, cytoskeleton, microtubule organizing center, centrosome. Generally nucleolar, but is translocated to the nucleoplasm in case of serum starvation or treatment with anticancer drugs. Has been found in the cytoplasm in patients with primary acute myelogenous leukemia (AML), but not with secondary AML. Can shuttle between cytoplasm and nucleus. Co- localizes with the methylated form of RPS10 in the granular component (GC) region of the nucleolus. Colocalized with nucleolin and APEX1 in nucleoli. Isoform 1 of NEK2 is required for its localization to the centrosome during mitosis.
  • Information by UniProt
  • Database links
  • Alternative names
    • 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
    • nucleophosmin nucleoplasmin family member 1 antibody
    • Nucleophosmin/nucleoplasmin family member 1 antibody
    • Numatrin antibody
    • OTTHUMP00000161024 antibody
    • OTTHUMP00000161025 antibody
    • OTTHUMP00000223397 antibody
    • OTTHUMP00000223398 antibody
    see all

Images

  • ab15440 detecting Nucleophosmin in interphase mouse primary embryonic fibroblasts. Cells were also counterstained with anti gamma tubulin antibody and DAPI in order to highlight the centrossomes and the nucleus (respectively). Secondary antibodies: goat anti-rabbit IgG conjugated to Alexa 488 (for ab15440) and goat anti-mouse IgG conjugated to Alexa 594 (for gamma-tubulin antibody).
  • ab15440 detecting Nucleophosmin in mitotic mouse primary embryonic fibroblasts. Cells were also counterstained with anti gamma tubulin antibody and DAPI in order to highlight the centrossomes and the nucleus (respectively). Secondary antibodies: goat anti-rabbit IgG conjugated to Alexa 488 (for ab15440) and goat anti-mouse IgG conjugated to Alexa 594 (for gamma-tubulin antibody).
  • Western blot analysis of various cell lines (25ug/lane) labeling Neucleophosmin with ab15440 at 1/500. A HRP-conjugated goat anti-rabbit IgG (1/20000) was used as the secondary antibody.

  • ab15440 staining human purified nucleophosmin from concentrated secreted serum free medium. 15 µg of purified protein were loaded in to each lane.  The primary antibody was diluted 1/500 and incubated with the blot for 15 hours at 4°C. A HRP conjugated donkey anti-rabbit IgG antibody, diluted 1/5000, was used as the secondary.

    See Abreview

  • ab15440 (4µg/ml) staining nucleophosmin in human colon using an automated system (DAKO Autostainer Plus). Using this protocol there is nuclear staining.
    Sections were rehydrated and antigen retrieved with the Dako 3-in-1 AR buffer citrate pH 6.0 in a DAKO PT Link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 minutes. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 minutes and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.
  • Immunohistochemistry was performed on both normal and cancer biopsies of deparaffinized human kidney tissue. To expose target proteins, heat induced antigen retrieval was performed using 10mM sodium citrate (pH6.0) buffer, microwaved for 8-15 minutes. Following antigen retrieval tissues were blocked in 3% BSA-PBS for 30 minutes at room temperature. Tissues were then probed at a dilution of 1/500 with a rabbit polyclonal antibody recognizing Nucleophosmin (ab15440) or without primary antibody (negative control) overnight at 4°C in a humidified chamber. Tissues were washed extensively with PBST and endogenous peroxidase activity was quenched with a peroxidase suppressor. Detection was performed using a biotin-conjugated secondary antibody and SA-HRP, followed by colorimetric detection using DAB. Tissues were counterstained with hematoxylin and prepped for mounting.

References

This product has been referenced in:
  • Holley CL  et al. Cytosolic accumulation of small nucleolar RNAs (snoRNAs) is dynamically regulated by NADPH oxidase. J Biol Chem 290:11741-8 (2015). WB ; Rat . Read more (PubMed: 25792744) »
  • Mayor R  et al. Genome distribution of replication-independent histone H1 variants shows H1.0 associated with nucleolar domains and H1X associated with RNA polymerase II-enriched regions. J Biol Chem 290:7474-91 (2015). WB ; Human . Read more (PubMed: 25645921) »
See all 17 Publications for this product

Customer reviews and Q&As

1-9 of 9 Abreviews or Q&A

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - whole cell (Saos)
Gel Running Conditions
Reduced Denaturing
Loading amount
20 µg
Specification
Saos
Blocking step
Milk as blocking agent for 3 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 24°C

Abcam user community

Verified customer

Submitted Feb 17 2016

Answer

Thank you for your email.

The answer you have provided to customer is very well structured. Perhaps you could also tell customer, that the information or data Abcam has about these products is already on the datasheet, they do not hold any more data than this.

We also haven't tested these antibodies in brain tissue sections infact it is not feasible for us to test every antibody in each body tissue however as the target is expressed in brain so the antibodies are fully guaranteed to work

Please forward the following link to customer (http://www.proteinatlas.org/ENSG00000181163/normal). It shows the target is expressed in brain.

I hope this information will be helpful.

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Purified protein (Concentrated secreted serum free medium)
Loading amount
15 µg
Specification
Concentrated secreted serum free medium
Gel Running Conditions
Reduced Denaturing (10%)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C

Dr. Aparna Mitra

Verified customer

Submitted Jul 11 2008

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Mouse Cell lysate - nuclear (Mouse embryonic stem cells)
Loading amount
20 µg
Specification
Mouse embryonic stem cells
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C

Abcam user community

Verified customer

Submitted Dec 05 2007

Answer

Thank you for your enquiry. You can do the experiment that you are describing but the primary antibodies must be of different isotypes. I have seen this experiment work best when you have one mouse antibody that is IgG and another that is IgM, and then you use an anti-IgG secondary and then an anti-IgM secondary. I do not believe there are any secondary antibodies that are specific for IgG1 versus IgG2, so you would have to do IgG versus IgM. I hope this information helps, please do not hesitate to contact us if you need any more advice or information.

Read More

Answer

I'm sorry to hear you are having a problem with ab15440. Thank you for your patience in waiting for my reply. It is difficult for me to determine at this point if the lack of staining is due to the primary or secondary antibody, or if the protocol could be adjusted to have success with the antibody. We have not had any complaints about the secondary or primary so it is difficult to determine. Since ab15440 is working in Western blot I am wondering if either the protocol needs to be adjusted or if the secondary is causing a problem for you. Immunofluorescence was added as an application because of the following review that a customer submitted: Sample Mouse Cell (embryonic stem cells) Application Immunocytochemistry/Immunofluorescence Blocking step: Serum as blocking agent for 2 hour(s) 0 minute(s) · Concentration: 10% Fixation: Paraformaldehyde Antigen retrieval step: None Incubation time: 12 hour(s) 0 minute(s) Dilution 1/100 Rating Good - I would definitely use it again Secondary antibody Name: Non-Abcam one was used: Alexa 568 Conjugation: do not use Dilution: 1/1000 The only major differences are that the review customer fixed the cells in paraformaldehyde, the primary incubation was overnight at 4 degrees, and the secondary was different. I would like to suggest the following modifications to your protocol: 1) Please use paraformaldehyde as a fixative instead of methanol:acetone as antibodies can be sensitive to the fixation time and method, 2) More importantly, please incubate in primary overnight at 4 degrees if you have not already done so, 3) Please confirm whether you have used the secondary with another primary to know that the secondary is giving you good results. Alternatively, please try a different secondary in IF so we will know if the secondary is causing the problems for you. Please let me know if this helps and do not hesitate to contact us for further advice.

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (embryonic stem cells)
Specification
embryonic stem cells
Fixative
Paraformaldehyde
Blocking step
Serum as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 10%

Abcam user community

Verified customer

Submitted Mar 01 2006

Answer

Thank you for your enquiry. Our recommended positive control for our Nucleophosmin antibody (ab15440) is a HeLa or mouse fibroblast extract. I would recommend our HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate (ab7898). I hope this information helps, please do not hesitate to contact me should you require further assistance.

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Mouse Cell lysate - whole cell (Primary hepatocytes)
Specification
Primary hepatocytes
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 10

Ms. Louise Mary Treanor

Verified customer

Submitted Nov 14 2005

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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