p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free

Novus Biologicals | Catalog # NB200-174

Novus Biologicals
Loading...

Key Product Details

Validated by

Knockout/Knockdown

Species Reactivity

Validated:

Human, Mouse, Golden Syrian Hamster (Negative)

Cited:

Human, Mouse

Applications

Validated:

Knockout Validated, Immunohistochemistry, Immunohistochemistry-Paraffin, Immunohistochemistry-Frozen, Western Blot, Flow Cytometry, Flow (Intracellular), Immunocytochemistry/ Immunofluorescence, Immunoprecipitation, CyTOF-ready

Cited:

Immunohistochemistry-Paraffin, Immunohistochemistry-Frozen, Western Blot, Flow Cytometry, Immunocytochemistry/ Immunofluorescence, IF/IHC

Label

Unconjugated

Antibody Source

Monoclonal Rat IgG2B Clone # 5-C3-1

Format

BSA Free
Loading...

Product Specifications

Immunogen

A synthetic peptide containing amino acids 54-75 of the murine p19ARF/CDKN2A. [UniProt# Q64364]

Epitope

The epitope recognized is between amino acids 62-75 of murine p19ARF/CDKN2A.

Reactivity Notes

Human reactivity reported in scientific literature (PMID: 27177758). The antibody does not react with Golden Syrian hamster. Armenian hamster has not been tested.

Clonality

Monoclonal

Host

Rat

Isotype

IgG2B

Theoretical MW

19 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Scientific Data Images for p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free

Western Blot: p19ARF/CDKN2A Antibody (5-C3-1)BSA Free [NB200-174]

Western Blot: p19ARF/CDKN2A Antibody (5-C3-1)BSA Free [NB200-174]

Western Blot: p19ARF/CDKN2A Antibody (5-C3-1) [NB200-174] - Whole cell protein from mouse Neuro2A and MEF cells was separated on a 4-20% gel by SDS-PAGE, transferred to 0.2 um PVDF membrane and blocked in 5% non-fat milk in TBST. The membrane was probed with 2.0 ug/mL anti-p19ARF in block buffer and detected with an anti-mouse HRP secondary antibody using chemiluminescence.
Immunohistochemistry: p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free [NB200-174]

Immunohistochemistry: p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free [NB200-174]

Immunohistochemistry: p19ARF/CDKN2A Antibody (5-C3-1) [NB200-174] - Staining of sections from a wild type mouse testis taken at P11 were probed with NB200-174 (green). Sections were stained with DAPI to reveal the position of cell nuclei (blue).
Flow (Intracellular): p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free [NB200-174]

Flow (Intracellular): p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free [NB200-174]

Flow (Intracellular): p19ARF/CDKN2A Antibody (5-C3-1) [NB200-174] - An intracellular stain was performed on HEK293 cells with p14ARF (5-C3-1) antibody NB200-174 (blue) and a matched isotype control MAB0061 (orange) [A]. An intracellular stain was also performed on THP-1 (known homozygous deletion of gene, PMID: 19635138) cells to ensure antibody specificity [B]. Cells were incubated in an antibody dilution of 1 ug/mL for 30 minutes at room temperature, followed by Goat Anti-Rat F(ab)2 IgG (H+L) APC-conjugated Antibody (F0113, R&D Systems). *Note: This antibody recognizes both human (p14ARF) and mouse (p19ARF) versions of the protein.
Western Blot: p19ARF/CDKN2A Antibody (5-C3-1)BSA Free [NB200-174]

Western Blot: p19ARF/CDKN2A Antibody (5-C3-1)BSA Free [NB200-174]

p19ARF-CDKN2A-Antibody-5-C3-1-Western-Blot-NB200-174-img0012.jpg
Western Blot: p19ARF/CDKN2A Antibody (5-C3-1)BSA Free [NB200-174]

Western Blot: p19ARF/CDKN2A Antibody (5-C3-1)BSA Free [NB200-174]

Western Blot: p19ARF/CDKN2A Antibody (5-C3-1) [NB200-174] - Analysis of endogenous p19Arf. Total cell lysates (25 ug) from NIH3T3 cells, which have deleted the Arf gene (Lanes 1) and from wild type mouse embryo fibroblasts (MEFs) at passage 6, which express p19Arf (Lanes 2), were resolved by SDS-PAGE.
Knockout Validated: p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free [NB200-174]

Immunocytochemistry/ Immunofluorescence: p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free [NB200-174]

Immunocytochemistry/ Immunofluorescence: p19ARF/CDKN2A Antibody (5-C3-1) [NB200-174] - Detection of p19Arf. Wild type mouse embryo fibroblasts (MEF) at passage 5 (panels 1, 2) and NIH3T3 cells, which have deleted the Arf gene (panels 3, 4) were probed with NB200-17 and nuclear stain (overlay in panels 2 and 4).
p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free

Western Blot: p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free [NB200-174] -

Western Blot: p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free [NB200-174] - Phosphorylation of NPM-Ser48 regulates the localization of NPM & p19A(A) Npm−/−, p53−/−double null MEF were infected with pBabe retrovirus expressing FLAG-tagged-myr-AKT1 in combination with NPM-WT or NPM-S48A as indicated. Cells were fixed & stained with DAPI & anti-NPM (left) or anti-phospho-S48-NPM (pS48-NPM). (B) NPM immunoprecipitates & whole cell lysates from Npm−/−;p53−/− MEFs expressing human NPM or NPM-S48A were probed with the indicated antibodies.(C) Graph, quantification of p19ARF staining intensity in confocal images using ImageJ. (D) Npm−/−, p53−/−double null MEF were infected with pBABE retrovirus expressing FLAG-tagged-myr-AKT1 in combination with NPM-WT, NPM-S48A or S48E as indicated. Cells were fixed & stained with DAPI, anti-NPM & anti p19ARF. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/25071014), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free

Western Blot: p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free [NB200-174] -

Western Blot: p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free [NB200-174] - Inhibition of AKT promotes enhanced MDM2 activity via the increased association between NPM & p14ARF(A) Npm−/−, p53−/−double null MEF were infected with pBABE retrovirus empty vector & pBABE expressing FLAG-tagged-NPM-WT, NPM-S48A or S48E as indicated. Immunopurification of NPM was done by pulling down with the Flag tag (middle panel) followed by elution of complexes by the Flag peptide & subsequent immunopurification of endogenous MDM2 (lower panel). (B) Nuclear immunoprecipitates of MDM2 from T24 cells treated with MK-2206 (5 μM, 24 hrs). Immunoprecipitates & lysates were blotted with the indicated antibodies. (C) T24 cells were treated with MK-2206 (5 μM) as indicated. p14ARF was immunoprecipitated from whole cell lysates & nuclear extracts & the association with NPM & MDM2 determined by western blot. Immunoprecipitates & lysates were blotted with the indicated antibodies. (D) MDM2 & (E) p53 ubiquinitation assay in H1299 cells transfected with wild type p53, HA-tagged ubiquitin & treated for 16 hrs with DMSO, MK-2206 (5 μM) or Nutlin3A (5 μM) as indicated. Immunoprecipitates & whole cell lysates were probed with the indicated antibodies. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/25071014), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free

Western Blot: p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free [NB200-174] -

Western Blot: p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free [NB200-174] - Loss of P19Arf enhances tumor growth & primary lung fibroblasts induce p19Arf in response to leucine deprivation (LD). (A) Graph of sarcoma (SKPY) tumor volume on the indicated days. SKPY cells were injected into the flank of WT & p19Arf-null mice. Tumor volume was measured by caliper. N=7 mice/WT cohort & N=15 mice/p19Arf−/− cohort. (B) Western blot for p19Arf protein expression in WT & Arf-null fibroblasts after LD for the indicated days. Densitometric quantification of p19Arf protein expression relative to beta -tubulin is shown on right. (C) qPCR for p19Arf mRNA in WT or p19Arf−/− fibroblasts during LD for the indicated days. (D) Representative immunofluorescence images for P19Arf subcellular localization on the indicated days after LD in WT or p19Arf−/− ALFs. Quantification of nuclear p19Arf is shown in graph on right. (E) Western blot analysis of phospho-eIF2 alpha (P-eIF2 alpha ) & total eIF2 alpha expression in WT & p19Arf-null murine ALFs during LD for the indicated days. Graph quantifies intensity of P-eIF2 alpha expression relative to day 0. beta -Tubulin used as loading control. (F) Western blot analysis of P-eIF2 alpha expression in WT & p19Arf−/− ALFs upon treatment with a GCN2 inhibitor (GCN2-IN-1; A-92 1 µM in DMSO) during overnight LD. Actin used as a loading control. Quantification of P-eIF2 alpha expression relative to total eIF2 alpha. N=3; *, P<0.05; ns, not significant. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/34994382), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free

Immunocytochemistry/ Immunofluorescence: p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free [NB200-174] -

Immunocytochemistry/ Immunofluorescence: p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free [NB200-174] - Phosphorylation of NPM-Ser48 regulates the localization of NPM & p19A(A) Npm−/−, p53−/−double null MEF were infected with pBabe retrovirus expressing FLAG-tagged-myr-AKT1 in combination with NPM-WT or NPM-S48A as indicated. Cells were fixed & stained with DAPI & anti-NPM (left) or anti-phospho-S48-NPM (pS48-NPM). (B) NPM immunoprecipitates & whole cell lysates from Npm−/−;p53−/− MEFs expressing human NPM or NPM-S48A were probed with the indicated antibodies.(C) Graph, quantification of p19ARF staining intensity in confocal images using ImageJ. (D) Npm−/−, p53−/−double null MEF were infected with pBABE retrovirus expressing FLAG-tagged-myr-AKT1 in combination with NPM-WT, NPM-S48A or S48E as indicated. Cells were fixed & stained with DAPI, anti-NPM & anti p19ARF. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/25071014), licensed under a CC-BY license. Not internally tested by Novus Biologicals.
p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free

Flow Cytometry: p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free

Flow Cytometry: p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free - Conducted the antibody trial with two different dilutions of p19ARF/CDKN2A Antibody (5-C3-1) [PE] (Catalog # NB200-174PE) (1:500 and 1:100) and compared them to an IgG control which was also PE labelled. Image from a verified customer review.

Applications for p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free

Application
Recommended Usage

Flow Cytometry

1 ug/mL

Immunocytochemistry/ Immunofluorescence

1:500-1:1000

Immunohistochemistry

5 u/gml

Immunohistochemistry-Frozen

5 ug/ml

Immunohistochemistry-Paraffin

reported in scientific literature (PMID 27177758)

Immunoprecipitation

1:10-1:500

Western Blot

1:500-1:1000
Application Notes
In Western blot a band can be seen at ~19 kDa. The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Flow Cytometry Panel Builder

Bio-Techne Knows Flow Cytometry

Save time and reduce costly mistakes by quickly finding compatible reagents using the Panel Builder Tool.

Advanced Features

  • Spectra Viewer - Custom analysis of spectra from multiple fluorochromes
  • Spillover Popups - Visualize the spectra of individual fluorochromes
  • Antigen Density Selector - Match fluorochrome brightness with antigen density
Build Your Panel Now

Formulation, Preparation, and Storage

Purification

Protein A purified

Formulation

PBS

Format

BSA Free

Preservative

0.02% Sodium Azide

Concentration

1.0 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.

Background: p19ARF/CDKN2A

ARF (alternative reading frame; p14ARF in human, p19ARF in mouse) is a tumor suppressor that serves as a sensor of hyper-proliferative signals, resulting in p53-dependent growth arrest and apoptotic cell death. ARF interacts with several proteins including COMMD1, CDKN2AIP, TBRG1, E4F1, C1QBP etc. and can bind to BCL6, E2F1, HUWE1, MDM2, MYC, NPM1/B23, TOP1/TOPOI as well as UBE2I/UBC9. ARF has the capability of inducing cell cycle arrest in G1 as well as G2 phases. While ARF is not expressed at significant levels in most normal tissues, oncogene activation triggers ARF overexpression, resulting in inhibition of MDM2 ubiquitin ligase and stabilization of p53. It is induced by progesterone as well as Ras/DMTF1, and is downregulated through NFkB p65 subunit (RELA). Inhibition of the p53 pathway, most commonly via mutations in p53 itself, inactivation of ARF, or amplification of MDM2 has been proposed to be a crucial step in pathogenesis of most human malignancies. Moreover, mice with null ARF have been shown to develop tumors rapidly and to display delayed mammary gland involution.

Alternate Names

alternative reading frame, ARF, ARF-INK4a, CDK4I, CDKN2A, Cyclin-dependent kinase 4 inhibitor A, cyclin-dependent kinase inhibitor 2A, cyclin-dependent kinase inhibitor 2A (p16, inhibits CDK4), cyclin-dependent kinase inhibitor 2A, isoform 3, cyclin-dependent kinase inhibitor 2A, isoforms 1/2, cyclin-dependent kinase inhibitor protein, INK4, INK4a, Ink4a/Arf, INK4a-ARF, mitochondrial smARF', MTS1, p14, p14ARF, P16, p16(INK4a), p16-INK4, P16INK4A, p16-INK4a, p19, p19, p19ARF, Pctr1

Gene Symbol

Cdkn2a

Additional p19ARF/CDKN2A Products

Product Documents for p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free

Certificate of Analysis

To download a Certificate of Analysis, please enter a lot or batch number in the search box below.

Product Specific Notices for p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Citations for p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free

Customer Reviews for p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free

There are currently no reviews for this product. Be the first to review p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free and earn rewards!

Have you used p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free?

Submit a review and receive an Amazon gift card!

$25/€18/£15/$25CAN/¥2500 Yen for a review with an image

$10/€7/£6/$10CAN/¥1110 Yen for a review without an image

Submit a review
Amazon Gift Card

Protocols

View specific protocols for p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free (NB200-174):


Immunoprecipitation

1. Total cell lysates (0.25 mg) from NIH3T3 cells which have deleted the Arf gene and from NIH3T3 cells expressing an HA-tagged version of p19Arf were incubated with control rat IgG or anti-p19Arf (NB 200-174) and protein G- Sepharose.
2. Precipitated proteins were resolved by SDS-PAGE and analysed by Western blotting using an anti-HA antibody. Unprecipitated total cell lysates (25 ug, equivalent to 10% of immunoprecipitation) from Arf null NIH3T3 cells and HA- p19Arf expressing cells were also analysed by Western blotting, as input controls.

Western Blot Protocol

1. Perform SDS-PAGE on samples to be analyzed, loading 10-25 ug of total protein per lane.
2. Transfer proteins to PVDF membrane according to the instructions provided by the manufacturer of the membrane and transfer apparatus.
3. Stain the membrane with Ponceau S (or similar product) to assess transfer success, and mark molecular weight standards where appropriate.
4. Rinse the blot TBS -0.05% Tween 20 (TBST).
5. Block the membrane in 5% Non-fat milk in TBST (blocking buffer) for at least 1 hour.
6. Wash the membrane in TBST three times for 10 minutes each.
7. Dilute primary antibody in blocking buffer and incubate overnight at 4C with gentle rocking.
8. Wash the membrane in TBST three times for 10 minutes each.
9. Incubate the membrane in diluted HRP conjugated secondary antibody in blocking buffer (as per manufacturer's instructions) for 1 hour at room temperature.
10. Wash the blot in TBST three times for 10 minutes each (this step can be repeated as required to reduce background).
11. Apply the detection reagent of choice in accordance with the manufacturer's instructions.

Find general support by application which include: protocols, troubleshooting, illustrated assays, videos and webinars.

FAQs for p19ARF/CDKN2A Antibody (5-C3-1) - BSA Free

Showing  1 - 1 of 1 FAQ Showing All
  • Q: We would like to confirm if the immunogen sequence is a synthetic peptide containing amino acids 54-75 (RRGPHRNPGPGDDDGQRSRSSS) of the murine p19ARF. And also, the epitope recognized is between amino acids 62-75 of murine p19ARF.

    A:

    I can confirm that the immunogen sequence for our product NB200-174 is aa 54-75 of the mouse Cdkn2a (also known as p19ARF) protein (Uniprot Site). The epitope recognized by this protein is aa 62-75.

Showing  1 - 1 of 1 FAQ Showing All
View all FAQs for Antibodies
Loading...