Overview

  • Product name
    Anti-Fibrillarin antibody [38F3]
    See all Fibrillarin primary antibodies
  • Description
    Mouse monoclonal [38F3] to Fibrillarin
  • Specificity
    This clone was selected because it stains a single ~34kDa band on western blotting and shows a clear and strong punctate staining of yeast nuclei. It can therefore be used to identify nucleoli immunocytochemically. ab4566 was raised against yeast nuclear preps and the immunogen was identified as Nop1p, the yeast homolog of fibrillarin. Due to high aa homology the antibody should work with any specie possessing a nucleus, however this has not been tested.
  • Tested applications
    Suitable for: Flow Cyt, IHC-P, ICC/IF, IHC-Fr, WB, ICCmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Chicken, Human, Arabidopsis thaliana, Caenorhabditis elegans, Drosophila melanogaster, Schizosaccharomyces pombe, Zebrafish, African green monkey, Opossum
    Predicted to work with: Plants
  • Immunogen

    Tissue, cells or virus corresponding to Saccharomyces cerevisiae Fibrillarin.

    Yeast nuclear preparation (S. cerevisiae). Hybridomas were screened by immunofluorescence on yeast cells and by western blotting on yeast protein homogenates (S. cerevisiae).

  • Positive control
    • ICC: Chicken embryonic fibroblast (CEF) cells, HEK 293 cells, SH-SY5Y cells
  • General notes

    Pfam number: PF01269. A reference below describes the characterization of D77, an antibody very similar but not identical to ab4566.

    Gives a much weaker signal in western blot compared to ab218846.

Properties

Applications

Our Abpromise guarantee covers the use of ab4566 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
Flow Cyt 1/20.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

IHC-P Use at an assay dependent concentration. PubMed: 18528333
ICC/IF 1/1000 - 1/5000.

For IF of mammalian cells 1/500.

ab4566 is sensitive to aldehyde. Use a mild formalin fixation or acetone or methanol fixation as the target is nuclear.

IHC-Fr Use at an assay dependent concentration. PubMed: 18559082
WB 1/2000 - 1/10000. Detects a band of approximately 34 kDa. 1/2000 (cell lysates) - 1/10000 (nuclear fractions)(ECL). For other (non-ECL) western detection methods, 1/1000 - 1/5000. To detect mammalian fibrillarin on western blots by ECL, 1/500.
ICC Use at an assay dependent concentration.

ab4566 is sensitive to aldehyde. Use a mild formalin fixation or acetone or methanol fixation as the target is nuclear.

Target

  • Function
    S-adenosyl-L-methionine-dependent methyltransferase that has the ability to methylate both RNAs and proteins. Involved in pre-rRNA processing by catalyzing the site-specific 2'-hydroxyl methylation of ribose moieties in pre-ribosomal RNA. Site specificity is provided by a guide RNA that base pairs with the substrate. Methylation occurs at a characteristic distance from the sequence involved in base pairing with the guide RNA. Also acts as a protein methyltransferase by mediating methylation of 'Gln-105' of histone H2A (H2AQ104me), a modification that impairs binding of the FACT complex and is specifically present at 35S ribosomal DNA locus (PubMed:24352239).
  • Sequence similarities
    Belongs to the methyltransferase superfamily. Fibrillarin family.
  • Post-translational
    modifications
    By homology to other fibrillarins, some or all of the N-terminal domain arginines are modified to asymmetric dimethylarginine (DMA).
  • Cellular localization
    Nucleus, nucleolus. Fibrillar region of the nucleolus.
  • Information by UniProt
  • Database links
    see all
  • Alternative names
    • 34 kD nucleolar scleroderma antigen antibody
    • 34 kDa nucleolar scleroderma antigen antibody
    • Fbl antibody
    • FBRL_HUMAN antibody
    • FIB antibody
    • FIB1 antibody
    • FLRN antibody
    • Histone-glutamine methyltransferase antibody
    • Nop1p antibody
    • RNA U3 small nucleolar interacting protein 1 antibody
    • RNU3IP1 antibody
    • rRNA 2' O methyltransferase fibrillarin antibody
    • rRNA 2'-O-methyltransferase fibrillarin antibody
    see all

Images

  • Rat neurons and glial stained with mouse monoclonal to Fibrillarin (green) and with chicken antibody to neurofilament NF-H (red). Cells were counterstained with a fluorescent DNA probe (blue). Nuclear DNA is revealed with Hoechst dye (blue). Cultures were processed using our standard fixation and staining procedure (in protocol section).

  • Human neuroblastoma line SH-SY5Y stained with mouse monoclonal to Fibrillarin (green) and with chicken antibody to neurofilament NF-H (red) and counterstained with a fluorescent DNA probe (blue). Nuclear DNA is revealed with Hoechst dye (blue). The NF-H antibody was used at a dilution of 1/100000 and the fibrillarin monoclonal at 1/1000. Cultures were processed using standard fixation and staining procedure (in protocol section).

  • Image courtesy of an anonymous Abreview.

    Mouse embryonic fibroblast fractionation.
    Cytopl - cytoplasmic fraction.
    Nucl - nuclear fraction.
    20 µg of each loaded.

    ab4566 used at a 1/2000 dilution.
    The secondary used was an Alexa-Fluor 680 conjugated goat anti-mouse polyclonal used at a 1/10000 dilution.

    See Abreview

  • All lanes : Anti-Fibrillarin antibody [38F3] (ab4566) at 1/2000 dilution (incubated for 1 hour, diluted with Blocking buffer 1:1 in PBS+0.1%Tween)

    Lane 1 : cytoplasmic protein fraction of HeLa cells with LI-COR® Odyssey® Blocking Buffer, 45 minutes at room temperature at 50 %
    Lane 2 : nuclear protein fraction of HeLa cells with LI-COR® Odyssey® Blocking Buffer, 45 minutes at room temperature at 50 %

    Lysates/proteins at 20 µg per lane.

    Secondary
    AlexaFluor 680 goat anti-mouse at 1/10000 dilution

    Performed under reducing conditions.

    Additional bands at : 34 kDa. We are unsure as to the identity of these extra bands.

    Exposure time : 1 second

    Image is courtesy of Dr Svetlana Khoronenkova

    See Abreview

  • Overlay histogram showing HEK293 cells stained with ab4566 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab4566, 1/20 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1](ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.

  • ab staining Fibrillarin in Human melanoma A7 cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with paraformaldehyde, permeabilized with 0.1% Triton and blocked with 1% BSA for 1 hour at room temperature. Samples were incubated with primary antibody (1/500 in 1% BSA) for 24 hours at 4°C. A FITC-conjugated Goat anti-mouse polyclonal (1/200) was used as the secondary antibody.

    See Abreview

  • High magnification view of human Hek293 cell nuclei stained with mouse monoclonal to fibrillarin (green), counterstained with a fluorescent DNA probe (blue). Nuclear DNA is revealed with Hoechst dye (blue). Cultures were processed using our standard fixation and staining procedure (in protocol section).

References

This product has been referenced in:
  • Krejcí J  et al. Neural Differentiation in HDAC1-Depleted Cells Is Accompanied by Coilin Downregulation and the Accumulation of Cajal Bodies in Nucleoli. Stem Cells Int 2017:1021240 (2017). IHC ; Mouse . Read more (PubMed: 28337219) »
  • Shekari F  et al. Data for whole and mitochondrial proteome of human embryonic stem cells. Data Brief 13:371-376 (2017). WB ; Human . Read more (PubMed: 28664172) »

See all 91 Publications for this product

Customer reviews and Q&As

Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (Embryonic Heart)
Permeabilization
Yes - Tween-20
Specification
Embryonic Heart
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Fixative
Paraformaldehyde
Username

Abcam user community

Verified customer

Submitted Oct 21 2016

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Mouse Tissue lysate - whole (Heart)
Gel Running Conditions
Reduced Denaturing (4-12%)
Loading amount
30 µg
Specification
Heart
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Abcam user community

Verified customer

Submitted Oct 05 2016

Application
Immunoprecipitation
Immuno-precipitation step
Protein A/G
Sample
Human Cell lysate - whole cell (HeLa cells)
Specification
HeLa cells
Total protein in input
30 µg
Username

Abcam user community

Verified customer

Submitted Jan 15 2015

Application
Immunocytochemistry/ Immunofluorescence
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: RT°C
Sample
Human Cell (Human primary fibroblasts)
Specification
Human primary fibroblasts
Permeabilization
Yes - 0.2% Triton-X100
Fixative
Paraformaldehyde
Username

Abcam user community

Verified customer

Submitted Dec 17 2013

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Expiration date: 21 March 2012


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The anti-Fibrillarin antibody [38F3] is a tissue culture supernatant and is not purified. Other proteins will be within the solution and this can give it an off white colour. ...

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Since you can see expected positivity along with what appears to be non specifici cytosolic staining, I would suggest diluting the primary 2 - 4 fold, which will hopefully reduce non specific background while retaining specificity.

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