Anti-Fibrillarin antibody - Nucleolar Marker (ab5821)

Overview

  • Product nameAnti-Fibrillarin antibody - Nucleolar Marker
    See all Fibrillarin primary antibodies
  • Description
    Rabbit polyclonal to Fibrillarin - Nucleolar Marker
  • SpecificityThis antibody detects a band at close to 34kDa in all species tested. The band can be completely blocked with the immuising peptide in all cases - this is very strong evidence that the antibody is recognising fibrillarin.
  • Tested applicationsSuitable for: IHC - Wholemount, ICC/IF, IP, IHC-P, WB, ICCmore details
  • Species reactivity
    Reacts with: Mouse, Human, Xenopus laevis, Drosophila melanogaster
    Does not react with: Rat
  • Positive control
    • This antibody gave a positive signal in the following whole cell lysates: HeLa; A431; Jurkat; 293T. This antibody also gave a positive signal in HeLa nuclear lysate.

Properties

Applications

Our Abpromise guarantee covers the use of ab5821 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 - Wholemount Use at an assay dependent concentration. PubMed: 22291607
ICC/IF Use at an assay dependent concentration.
IP Use at an assay dependent concentration. PubMed: 19430468
IHC-P 1/100.
WB Use a concentration of 1 µg/ml. Detects a band of approximately 39 kDa (predicted molecular weight: 34 kDa).
ICC 1/100.

Target

  • FunctionS-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 similaritiesBelongs 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 localizationNucleus, 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

Anti-Fibrillarin antibody - Nucleolar Marker images

  • ICC/IF image of ab5821 stained human HeLa cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab5821, 1µg/ml) for 1h at room temperature. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iTTM FX Signal Enhancer was used as the primary blocking agent, 5% BSA (in TBS-T) was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue). Alexa Fluor® 594 WGA was used to label plasma membranes (red).

  • All lanes : Anti-Fibrillarin antibody - Nucleolar Marker (ab5821) at 1 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 2 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate
    Lane 3 : Schneider L2 whole cell lysate (ab14893)

    Lysates/proteins at 20 µg per lane.

    Secondary
    Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) (ab65484) at 1/3000 dilution

    Performed under reducing conditions.

    Predicted band size : 34 kDa
    Observed band size : 37 kDa (why is the actual band size different from the predicted?)
    Additional bands at : 42 kDa,70 kDa. We are unsure as to the identity of these extra bands.

    Exposure time : 3 minutes
  • The cells were 100% methanol fixed (5 min) and then
    incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab37266, 1µg/ml, red) and (ab5821, 1µg/ml, blue) overnight at +4°C. The secondary antibodies were ab150115 Alexa Fluor® 647 goat anti-mouse IgG (H+L) used at 2µg/ml for 1h and ab175652 Alexa Fluor® 405 goat anti-rabbit IgG (H+L) used at 2µg/ml for 1h. 

  • Immunofluorescent imaging of human cells (U2OS) with ab5821 reveals highly specific localisation to the dense fibrillar component (DFC) of the nucleolus associated with the initial ribosomal RNA (rRNA) precursor. The nucleolar protein fibrillarin is located primarily in the DFC.  Blue is hoechst staining of the nucleus, green is ab5821 used at 1/100, merge image demonstrates exclusively nuclear localisation.  

    IF was performed with a standard paraformaldehyde technique (fixed in PBS buffered PFH 4% for 5 minutes, permeabilised with 0.5% triton-PBS for 5 minutes, blocked with 5% milk / 0.2% tween for one hour.  Primary antibody used at 1/100 in 5% milk / 0.2% TWEEN for one hour, secondary antibody Alexa 488 for 30 minutes.  All blocking and incubation steps carried out at 37 degrees C.

  • ab5821 staining human liver tissue sections by IHC-P.  Sections were formaldehyde fixed, permeabilized in Triton-X and subjected to heat mediated antigen retrieval prior to blocking with 6% BSA for 2 hours at 23°C. The primary antibody was diluted 1/100 and incubated with the sample for 16 hours at 5°C.  ab6717 was used as the secondary antibody.

    See Abreview

  • Immunohistochemical analysis of salivary glands from Drosophila melanogaster larvae, staining Fibrillarin with ab5821. Tissues were fixed in 4% paraformaldehyde/PBS and 0.3% Triton-X/PBS before incubation with primary antibody.

References for Anti-Fibrillarin antibody - Nucleolar Marker (ab5821)

This product has been referenced in:
  • Terrier O  et al. Nucleolin interacts with influenza A nucleoprotein and contributes to viral ribonucleoprotein complexes nuclear trafficking and efficient influenza viral replication. Sci Rep 6:29006 (2016). ICC/IF ; Human . Read more (PubMed: 27373907) »
  • Goudarzi KM  et al. mTOR inhibitors blunt the p53 response to nucleolar stress by regulating RPL11 and MDM2 levels. Cancer Biol Ther 15:1499-514 (2014). Human . Read more (PubMed: 25482947) »

See all 51 Publications for this product

Product Wall

Application Immunocytochemistry/ Immunofluorescence
Sample Monkey Cell (Kidney)
Permeabilization Yes - 0.1% triton x-100
Specification Kidney
Blocking step Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 37°C
Fixative Paraformaldehyde
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Submitted Jun 10 2016

Application Western blot
Sample Monkey Cell lysate - whole cell (Kidney)
Gel Running Conditions Non-reduced Non-Denaturing (Native)
Loading amount 30 µg
Specification Kidney
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 25°C
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Submitted May 04 2016

Application Immunohistochemistry (Frozen sections)
Sample Mouse Tissue sections (Brain)
Permeabilization Yes - 0.3% Triton x 100: 30 min
Specification Brain
Blocking step Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 37°C
Fixative Paraformaldehyde
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Submitted May 04 2016

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Mouse Cell (mouse embryonic stem cells (E14 mES))
Permeabilization Yes - 0.25% triton
Specification mouse embryonic stem cells (E14 mES)
Blocking step BSA as blocking agent for 20 minute(s) · Concentration: 0.5% · Temperature: RT°C
Fixative Paraformaldehyde
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Submitted Jan 05 2016

Application Western blot
Sample Mouse Tissue lysate - other (Brain)
Gel Running Conditions Reduced Denaturing (10)
Loading amount 20 µg
Specification Brain
Blocking step Odyssey Blocking Buffer (PBS) as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 45% · Temperature: 25°C
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Submitted Sep 18 2015

Application Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample Mouse Tissue sections (Brain)
Antigen retrieval step None
Permeabilization Yes - Tween-20
Specification Brain
Blocking step Heat-Inactivated Normal Donkey Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Fixative Paraformaldehyde
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Submitted Apr 27 2015

Application Western blot
Loading amount 50 µg
Gel Running Conditions Reduced Denaturing
Sample Xenopus laevis Cell lysate - nuclear (adult heart)
Specification adult heart
Blocking step Milk as blocking agent for 4 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 23°C
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Submitted Aug 20 2014

Application Immunocytochemistry
Blocking step Innovex Background buster as blocking agent for 30 minute(s) · Concentration: 100% · Temperature: 25°C
Sample Human Cultured Cells (Pancreatic cancer cell line)
Specification Pancreatic cancer cell line
Permeabilization Yes - 0.25% tritonx X-100
Fixative Formaldehyde
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Submitted May 02 2014

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Application Immunocytochemistry/ Immunofluorescence
Sample Human Cell (MRC-5)
Specification MRC-5
Fixative Paraformaldehyde
Permeabilization Yes - 0.2% Triton X-100
Blocking step BSA as blocking agent for 30 minute(s) · Concentration: 1% · Temperature: RT°C
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Submitted Feb 11 2013

Thank you for your response. Unfortunately, we do not have the pdf format of our on-line product datasheets yet but IT Department will consider implementing it. I have saved the on-line documents as word documents and attached the files ...

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