Anti-Fibrillarin抗体- Nucleolar Marker (ab5821)

概述

  • 产品名称
    Anti-Fibrillarin抗体- Nucleolar Marker
    参阅全部 Fibrillarin 一抗
  • 描述
    兔多克隆抗体to Fibrillarin - Nucleolar Marker
  • 特异性
    This 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.
  • 经测试应用
    适用于: IHC - Wholemount, ICC/IF, IP, IHC-P, WB, ICCmore details
  • 种属反应性
    与反应: Mouse, Human, Xenopus laevis, Drosophila melanogaster
    不与反应: Rat
  • 免疫原

    Synthetic peptide corresponding to Human Fibrillarin aa 1-100 (C terminal) conjugated to keyhole limpet haemocyanin.
    Database link: P22087
    (Peptide available as ab13745)

  • 阳性对照
    • 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. ICC-IF: Hela cells.

性能

应用

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.

应用 Ab评论 说明
IHC - Wholemount Use at an assay dependent concentration. PubMed: 22291607
ICC/IF Use a concentration of 0.1 - 1 µg/ml.
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.

靶标

  • 功能
    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).
  • 序列相似性
    Belongs to the methyltransferase superfamily. Fibrillarin family.
  • 翻译后修饰
    By homology to other fibrillarins, some or all of the N-terminal domain arginines are modified to asymmetric dimethylarginine (DMA).
  • 细胞定位
    Nucleus, nucleolus. Fibrillar region of the nucleolus.
  • Information by UniProt
  • 数据库链接
    see all
  • 别名
    • 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

图片

  • 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
  • ab5821 stained in Hela cells. Cells were fixed with 100% methanol (5 min) at room temperature and incubated with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% triton for 1h at room temperature to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody ab5821 at 0.1 µg/ml and ab7291 (Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control) at 1/1000 dilution overnight at +4°C. The secondary antibodies were ab150120 (pseudo-colored red) and ab150081 (colored green) used at 1 µg/ml for 1 hour at room temperature. DAPI was used to stain the cell nuclei (colored blue) at a concentration of 1.43 µM for 1hour at room temperature.

  • 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.

文献

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). WB ; Mouse . Read more (PubMed: 28337219) »
  • Martins T  et al. TGFß/Activin signalling is required for ribosome biogenesis and cell growth in Drosophila salivary glands. Open Biol 7:N/A (2017). Read more (PubMed: 28123053) »

See all 85 Publications for this product

客户评价及客户问答

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Mouse Tissue lysate - whole (Whole brain tissue lysate)
Gel Running Conditions
Non-reduced Non-Denaturing (Native) (3-8% Tris-Acetate)
Loading amount
15 µg
Specification
Whole brain tissue lysate
Blocking step
BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C
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Verified customer

提交于 Apr 10 2017

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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提交于 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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提交于 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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Verified customer

提交于 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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提交于 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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提交于 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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提交于 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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提交于 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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提交于 May 02 2014

Abcam guarantees this product to work in the species/application used in this Abreview.
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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提交于 Feb 11 2013

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