Overview

  • Product name
    Anti-Argonaute-2 antibody - ChIP Grade
    See all Argonaute-2 primary antibodies
  • Description
    Rabbit polyclonal to Argonaute-2 - ChIP Grade
  • Host species
    Rabbit
  • Tested applications
    Suitable for: IP, IHC-P, ChIP, RIP, WB, ICC/IFmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
    Predicted to work with: Rabbit, CowDoes not react with: Drosophila melanogaster
  • Immunogen

    Synthetic peptide corresponding to Argonaute-2 aa 350-450 conjugated to keyhole limpet haemocyanin.
    (Peptide available as ab32380)

  • Positive control
    • WB: HeLa whole cell lysate. IHC-P: Human breast cancer tissue.

Properties

Applications

Our Abpromise guarantee covers the use of ab32381 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
IP Use at an assay dependent concentration. PubMed: 24280866
IHC-P Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
ChIP Use at an assay dependent concentration. PubMed: 19696410
RIP Use at an assay dependent concentration.
WB Use a concentration of 2 µg/ml. Detects a band of approximately 87 kDa (predicted molecular weight: 97 kDa).
ICC/IF 1/200.

Target

  • Function
    Required for RNA-mediated gene silencing (RNAi) by the RNA-induced silencing complex (RISC). The 'minimal RISC' appears to include EIF2C2/AGO2 bound to a short guide RNA such as a microRNA (miRNA) or short interfering RNA (siRNA). These guide RNAs direct RISC to complementary mRNAs that are targets for RISC-mediated gene silencing. The precise mechanism of gene silencing depends on the degree of complementarity between the miRNA or siRNA and its target. Binding of RISC to a perfectly complementary mRNA generally results in silencing due to endonucleolytic cleavage of the mRNA specifically by EIF2C2/AGO2. Binding of RISC to a partially complementary mRNA results in silencing through inhibition of translation, and this is independent of endonuclease activity. May inhibit translation initiation by binding to the 7-methylguanosine cap, thereby preventing the recruitment of the translation initiation factor eIF4-E. May also inhibit translation initiation via interaction with EIF6, which itself binds to the 60S ribosomal subunit and prevents its association with the 40S ribosomal subunit. The inhibition of translational initiation leads to the accumulation of the affected mRNA in cytoplasmic processing bodies (P-bodies), where mRNA degradation may subsequently occur. In some cases RISC-mediated translational repression is also observed for miRNAs that perfectly match the 3' untranslated region (3'-UTR). Can also upregulate the translation of specific mRNAs under certain growth conditions. Binds to the AU element of the 3'-UTR of the TNF (TNF-alpha) mRNA and upregulates translation under conditions of serum starvation. Also required for transcriptional gene silencing (TGS), in which short RNAs known as antigene RNAs or agRNAs direct the transcriptional repression of complementary promoter regions.
  • Sequence similarities
    Belongs to the argonaute family. Ago subfamily.
    Contains 1 PAZ domain.
    Contains 1 Piwi domain.
  • Domain
    The Piwi domain may perform RNA cleavage by a mechanism similar to that of RNase H. However while RNase H utilizes a triad of Asp-Asp-Glu (DDE) for metal ion coordination, this protein appears to utilize a triad of Asp-Asp-His (DDH).
  • Post-translational
    modifications
    Hydroxylated. 4-hydroxylation appears to enhance protein stability but is not required for miRNA-binding or endonuclease activity.
  • Cellular localization
    Cytoplasm > P-body. Nucleus. Translational repression of mRNAs results in their recruitment to P-bodies. Translocation to the nucleus requires IMP8.
  • Information by UniProt
  • Database links
  • Alternative names
    • Ago 2 antibody
    • AGO2_HUMAN antibody
    • Argonaute 2 antibody
    • argonaute 2, RISC catalytic component antibody
    • Argonaute RISC catalytic component 2 antibody
    • Argonaute2 antibody
    • CTA-204B4.6 antibody
    • dAgo2 antibody
    • eIF 2C 2 antibody
    • eIF-2C 2 antibody
    • eIF2C 2 antibody
    • Eif2c2 antibody
    • Eukaryotic translation initiation factor 2C 2 antibody
    • Eukaryotic translation initiation factor 2C subunit 2 antibody
    • hAgo2 antibody
    • MGC3183 antibody
    • PAZ Piwi domain protein antibody
    • PPD antibody
    • Protein argonaute-2 antibody
    • Protein slicer antibody
    • Q10 antibody
    • Slicer protein antibody
    see all

Images

  • Immunohistochemical analysis of Argonaute2 (AGO2) in human tissues sections from slowly progressive, rapidly progressive, or normal biopsies.

    Representative images of slowly progressive (A–B, E–F), rapidly progressive (C–D, G–H), and normal (I–J) biopsies stained with IgG control (A, C, E, G, & I) and anti-AGO2 antibody (B, D, F, H, & J) are shown. Images A, B, C, & D were stained with antibody concentrations of 20 µg/ml and images E, F, G, H, I, & J were stained with an antibody concentration of 4 µg/ml. Sections were counterstained with hematoxylin. Protein expression stains brown in this procedure (original magnification: ×200).

    Panels A-D shown.

  • All lanes : Anti-Argonaute-2 antibody - ChIP Grade (ab32381) at 2 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
    Lane 2 : Jurkat whole cell lysate (ab7899)
    Lane 3 : HeLa (Human epithelial carcinoma cell line) Nuclear Lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : IRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/10000 dilution

    Performed under reducing conditions.

    Predicted band size: 97 kDa
    Observed band size: 87 kDa
    why is the actual band size different from the predicted?
    Additional bands at: 55 kDa. We are unsure as to the identity of these extra bands.



    The identification of the 55 kDa band is unclear but this band has also been observed in HeLa lysates in the Western Blot of ab5072, targeting Drosophila Ago2 / eIF2C2.

  • ab32381 (1/200) staining Ago2/ eIF2C2 in asynchronous HeLa (Human epithelial cell line from cervix adenocarcinoma) cells (green). Cells were fixed with paraformaldehyde, permeabilized with 0.5% TritonX100/ PBS and counterstained with DAPI in order to highlight the nucleus (red). Please refer to abreview for further experimental details.

    See Abreview

  • IHC image of Ago2/eIF2C2 staining in human breast cancer FFPE section, performed on a BondTM system using the standard protocol F.

    The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab32381, 5µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with hematoxylin and mounted with DPX.

  • RIP analysis of 368T1 murine lung cancer cells using ab32381 to bind Ago2 / eIF2C2.

    Cells were transfected with an empty vector (empty), a vector expressing wild-type Hmga2 (wt), a vector expressing a let-7 site mutated Hmga2 (m7), a vector expressing wild-type Hmga2 without a start codon (ATG wt), or a vector expressing a let-7 site mutated Hmga2 without a start codon (ATG m7). RIP was performed with 5 µg Ago2 / eIF2C2 or IgG antibody per 107 cells. Binding was detected using qRT-PCR.

    See Abreview

References

This product has been referenced in:
  • Liang W  et al. Knockdown of growth-arrest specific transcript 5 restores oxidized low-density lipoprotein-induced impaired autophagy flux via upregulating miR-26a in human endothelial cells. Eur J Pharmacol 843:154-161 (2019). Read more (PubMed: 30468731) »
  • Dong H  et al. Activation of LncRNA TINCR by H3K27 acetylation promotes Trastuzumab resistance and epithelial-mesenchymal transition by targeting MicroRNA-125b in breast Cancer. Mol Cancer 18:3 (2019). Read more (PubMed: 30621694) »
See all 69 Publications for this product

Customer reviews and Q&As

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1-10 of 11 Abreviews

Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (mouse muscle tissue)
Permeabilization
Yes - Triton X-100
Specification
mouse muscle tissue
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 0.1% · Temperature: 22°C
Fixative
Paraformaldehyde

Nesrin Sabha

Verified customer

Submitted Feb 28 2018

Application
RIP
Sample
Mouse Cell lysate - whole cell (murine lung cancer cells)
Specification
murine lung cancer cells
Type
RIP without cross-linking
Duration of cross-linking step: 0 second(s)
Detection step
Real-time PCR
Positive control
empty vector cells for miR-10a
Negative control
RIP with control IgG
Immuno-precipitation step
Protein G

Abcam user community

Verified customer

Submitted Nov 15 2012

Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (HeLa)
Specification
HeLa
Fixative
Paraformaldehyde
Permeabilization
Yes - 0.5 % Tritionx100
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 23°C

Abcam user community

Verified customer

Submitted Oct 05 2012

Application
Western blot
Sample
Mouse Cell lysate - whole cell (Mouse nueroblastoma N2A cell)
Loading amount
30 µg
Specification
Mouse nueroblastoma N2A cell
Gel Running Conditions
Reduced Denaturing (10%)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C

Abcam user community

Verified customer

Submitted Jul 03 2012

Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (saos-2 human osteosarcoma cell line)
Specification
saos-2 human osteosarcoma cell line
Fixative
Paraformaldehyde
Permeabilization
Yes - PBS TRITON 0.25%
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: RT°C

Abcam user community

Verified customer

Submitted Apr 19 2012

Application
CLIP
Sample
Human Cell lysate - whole cell (HEK 293T)
Specification
HEK 293T
Type
iCLIP
Irradiation Energy Level (mJ/cm2): 150
Wavelength (nm): 254
Positive control
Anti-hnRNP C antibody (sc-15386) bound protein A dynabeads, 0.1 mg/ml (antibody/beads)
Negative control
Protein A dynabeads without antibody bound
Immuno-precipitation step
Other - Protein A dynabeads

Abcam user community

Verified customer

Submitted Mar 21 2012

Application
Western blot
Sample
Human Cell lysate - whole cell (HeLa, HEK293)
Loading amount
40 µg
Specification
HeLa, HEK293
Gel Running Conditions
Reduced Denaturing (7.5)
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C

Abcam user community

Verified customer

Submitted Feb 18 2011

Application
Western blot
Sample
Human Cell lysate - whole cell (HeLa NIH-3T3 MEF GC-1)
Loading amount
20 µg
Specification
HeLa NIH-3T3 MEF GC-1
Gel Running Conditions
Reduced Denaturing (Precise protein gel 4-20%)
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 21°C

Mr. Antibody Solutions

Verified customer

Submitted Jun 16 2009

Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (HeLa)
Specification
HeLa
Fixative
Paraformaldehyde
Permeabilization
Yes - 0.5% TritonX100 in PBS

Dr. Kirk Mcmanus

Verified customer

Submitted May 28 2009

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (Brain)
Specification
Brain
Fixative
Formaldehyde
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Dako Target Retreival Solution
Permeabilization
No
Blocking step
5 minutes of peroxidase block then 10 minutes of protein block. These are ready-to-use reagents purchased from Dako as blocking agent for 15 minute(s) · Concentration: 100% · Temperature: 20°C

Mr. Antibody Solutions

Verified customer

Submitted Apr 07 2009

1-10 of 11 Abreviews

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