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Nucleolin Antibody

Catalog # NB600-241 | Novus Biologicals a Bio-Techne Brand
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NB600-241
NB600-241SS

Key Product Details

Species Reactivity

Human, Mouse, Rat, Plant

Applications

Chromatin Immunoprecipitation, Chromatin Immunoprecipitation (ChIP), Electron Microscopy, Immunocytochemistry/ Immunofluorescence, Immunohistochemistry, Immunohistochemistry Whole-Mount, Immunohistochemistry-Frozen, Immunohistochemistry-Paraffin, Immunomicroscopy, Knockdown Validated, Simple Western, Western Blot

Label

Unconjugated

Antibody Source

Polyclonal Rabbit IgG

Concentration

This product is unpurified. The exact concentration of antibody is not quantifiable.

Product Summary for Nucleolin Antibody

Immunogen

A fusion protein containing amino acids 284-709 of human Nucleolin. [UniProt# P19338]

Localization

Nucleoli

Clonality

Polyclonal

Host

Rabbit

Isotype

IgG

Theoretical MW

106 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 Nucleolin Antibody

Immunocytochemistry/Immunofluorescence: Nucleolin Antibody [NB600-241] - Analysis of Nucleolin MCF-7 breast cancer cells. Image courtsey of product review by Lacey Litchfield.
Simple Western: Nucleolin Antibody [NB600-241] - Image shows a specific band for Nucleolin in 0.5 mg/mL of HeLa lysate. This experiment was performed under reducing conditions using the 12-230 kDa separation system.
Immunocytochemistry/Immunofluorescence: Nucleolin Antibody [NB600-241] - Endogenous nuclear nucleolin-COUP-TFII interaction in MCF-7 and T47D cells.NE (200A ug protein) from MCF-7 cells. Immunofluorescent staining of endogenous COUP-TFII (green) and nucleolin (red) in the nuclei (Hoechst, blue) of MCF-7 cells. Merged images are shown at the right. Bar is 10A um. Image collected and cropped by Citeab from the following publication (Identification and characterization of nucleolin as a COUP-TFII coactivator of retinoic acid receptor transcription in breast cancer cells. PLoS One (2012) licensed under a CC-BY license.

Applications for Nucleolin Antibody

Application
Recommended Usage

Chromatin Immunoprecipitation

reported in scientific literature

Electron Microscopy

1:10-1:500

Immunocytochemistry/ Immunofluorescence

1:50-1:100

Immunohistochemistry

1:50-1:100

Immunohistochemistry Whole-Mount

reported in scientific literature (PMID 22483616)

Immunohistochemistry-Frozen

reported in scientific literature (PMID 30411850)

Immunohistochemistry-Paraffin

1:50-1:100

Simple Western

1:400

Western Blot

1:4000
Application Notes
In Western blot, a band at 1~106 kDa represents human Nucleolin. Heat mediated antigen retrieval is recommended for IHC-P.

In Simple Western only 10 - 15 uL of the recommended dilution is used per data point. Separated by Size-Wes, Sally Sue/Peggy Sue.
Please Note: Optimal dilutions of this antibody should be experimentally determined.

Reviewed Applications

Read 2 reviews rated 5 using NB600-241 in the following applications:

Published Applications

Read 25 publications using NB600-241 in the following applications:

Formulation, Preparation, and Storage

Purification

Unpurified

Formulation

Whole antisera

Preservative

0.1% Sodium Azide

Concentration

This product is unpurified. The exact concentration of antibody is not quantifiable.

Shipping

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

Stability & Storage

Aliquot and store at -20C or -80C. Avoid freeze-thaw cycles.

Background: Nucleolin

Nucleolin (NCL, also called Protein C23) is a major phosphoprotein present in nucleolus of growing eukaryotic cells which is implicated in the synthesis as well as maturation of ribosomes. Localized mainly to nucleolus, NCL may also be found in cytoplasmic mRNP granules containing untranslated mRNAs (mRNP granule complex contains - NCL, ACTB, ACTN4, DHX9, ERG, HNRNPA1, HNRNPA2B1, HNRNPAB, HNRNPD, HNRNPL, HNRNPR, HNRNPU, HSPA1, HSPA8, IGF2BP1, ILF2, ILF3, NCBP1, PABPC1, PABPC4, PABPN1, RPLP0, RPS3, RPS3A, RPS4X, RPS8, RPS9, SYNCRIP, TROVE2, YBX1 and untranslated mRNAs). NCL interacts with APTX, NSUN2, TERT, ERBB4, GZF1, GZF1, NVL and C1QBP, and is a part of SWAP complex (NPM1, NCL, PARP1 and SWAP70) as well as another complex containing NCL, HTATSF1/Tat-SF1, CDK9 and CCNT1, RNA polymerase II, SUPT5H. NCL may be found associated with intranucleolar chromatin and pre-ribosomal particles where it induces chromatin de-condensation via binding to histone H1 and play a key role in pre-rRNA transcription as well as ribosome assembly. NCL has also been suggested to implicate in the process of transcriptional elongation and it binds RNA oligonucleotides with 5'-UUAGGG-3' repeats more tightly than telomeric single-stranded DNA 5'-TTAGGG-3' repeats. Besides being essential to rRNA processing in nucleolus, NCL has several other functions such as transcription regulation, modulation of mRNA stability and DNA replication and recombination.

Alternate Names

C23, FLJ45706, FLJ59041, nucleolin, Protein C23

Entrez Gene IDs

4691 (Human); 17975 (Mouse)

Gene Symbol

NCL

UniProt

Product Documents for Nucleolin Antibody

Certificate of Analysis

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

Product Specific Notices for Nucleolin Antibody

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.

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Protocols

View specific protocols for Nucleolin Antibody (NB600-241):

[[URL:https://www.novusbio.com/products/nucleolin-antibody_nb600-241]][[Capti… Antibody]]
IHC-FFPE sections


I. Deparaffinization:

A. Treat slides with Xylene: 3 changes for 5 minutes each. Drain slides for 10 seconds between changes.

B. Treat slides with 100% Reagent Alcohol: 3 changes for 5 minutes each. Drain slides for 10 seconds between changes.


II. Quench Endogenous Peroxidase:

A. Place slides in peroxidase quenching solution: 15-30 minutes. To Prepare 200 ml of Quenching Solution: Add 3 ml of 30% Hydrogen Peroxide to 200 ml of Methanol.
Use within 4 hours of preparation

B. Place slides in distilled water: 2 changes for 2 minutes each.


III. Retrieve Epitopes:

A. Preheat Citrate Buffer. Place 200 ml of Citrate Buffer Working Solution into container, cover and place into steamer. Heat to 90-96 degrees Celsius.

B. Place rack of slides into hot Citrate Buffer for 20 minutes. Cover.

C. Carefully remove container with slides from steamer and cool on bench, uncovered, for 20 minutes.

D. Slowly add distilled water to further cool for 5 minutes.

E. Rinse slides with distilled water. 2 changes for 2 minutes each.


IV. Immunostaining Procedure:

A. Remove each slide from rack and circle tissue section with a hydrophobic barrier pen (e.g. Liquid Blocker-Super Pap Pen).

B. Flood slide with Wash Solution. Do not allow tissue sections to dry for the rest of the procedure.

C. Drain wash solution and apply 4 drops of Blocking Reagent to each slide and incubate for 15 minutes.

D. Drain Blocking Reagent (do not wash off the Blocking Reagent), apply 200 ul of Primary Antibody solution to each slide, and incubate for 1 hour.

E. Wash slides with Wash Solution: 3 changes for 5 minutes each.

F. Drain wash solution, apply 4 drops of Secondary antibody to each slide and incubate for 1 hour.

G. Wash slides with Wash Solution: 3 changes for 5 minutes each.

H. Drain wash solution, apply 4 drops of DAB Substrate to each slide and develop for 5-10 minutes. Check development with microscope.

I. Wash slides with Wash Solution: 3 changes for 5 minutes each.

J. Drain wash solution, apply 4 drops of Hematoxylin to each slide and stain for 1-3 minutes. Increase time if darker counterstaining is desired.

K. Wash slides with Wash Solution: 2-3 changes for 2 minutes each.

L. Drain wash solution and apply 4 drops of Bluing Solution to each slide for 1-2 minutes.

M. Rinse slides in distilled water.

N. Soak slides in 70% reagent alcohol: 3 minutes with intermittent agitation.

O. Soak slides in 95% reagent alcohol: 2 changes for 3 minutes each with intermittent agitation.

P. Soak slides in 100% reagent alcohol: 3 changes for 3 minutes each with intermittent agitation. Drain slides for 10 seconds between each change.

Q. Soak slides in Xylene: 3 changes for 3 minutes each with intermittent agitation. Drain slides for 10 seconds between each change.

R. Apply 2-3 drops of non-aqueous mounting media to each slide and mount coverslip.

S. Lay slides on a flat surface to dry prior to viewing under microscope.


NOTES:

-Use treated slides (e.g. HistoBond) to assure adherence of FFPE sections to slide.

-Prior to deparaffinization, heat slides overnight in a 60 degrees Celsius oven.

-All steps in which Xylene is used should be performed in a fume hood.

-For Epitope Retrieval, a microwave or pressure cooker may be substituted for the steamer method. Adjust times as necessary depending on conditions.

-For the initial IHC run with a new primary antibody, test tissues with and without Epitope Retrieval. In some instances, Epitope Retrieval may not be necessary.

-200 ul is the recommended maximum volume to apply to a slide for full coverage. Using more than 200 ul may allow solutions to wick off the slide and create drying artifacts. For small tissue sections less than 200 ul may be used.

-5 minutes of development with DAB Substrate should be sufficient. Do not develop for more than 10 minutes. If 5 minutes of development causes background staining, further dilution of the primary antibody may be necessary.

-Hematoxylin should produce a light nuclear counterstain so as not to obscure the DAB staining. Counterstain for 1-1.5 minutes for nuclear antigens. Counterstain for 2-3 minutes for cytoplasmic and membranous antigens. If darker counterstaining is desired increase time (up to 10 minutes).

FAQs for Nucleolin Antibody

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  • Q: Have you tested NB600-241 on nucleus fraction or cytoplasmic fraction? Do you have any publication or data could support the signal only appear on nucleus?

    A: Yes, it has been tested in nuclear extracts. Furthermore, it appears that a customer detected nucleolin in both the cytomplasm and the nucleus. Nucleolin appears to reside in both the cytoplasm and the nucleus (UniProt P19338) so it would be impossible to create an antibody that only detects nucleolin in just the nucleus. Our 10+ publications using this antibody can also be found on our website. These papers may provide further insight beyond our internal testing.

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