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    sirt1-antibody-e104-ab32441.pdf

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Cell Biology Apoptosis Intracellular p53 Pathway
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RecombinantRabMAb

Recombinant Anti-SIRT1 antibody [E104] (ab32441)

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Reviews (13)Q&A (15)References (126)

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Western blot - Anti-SIRT1 antibody [E104] (ab32441)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIRT1 antibody [E104] (ab32441)
  • Immunocytochemistry/ Immunofluorescence - Anti-SIRT1 antibody [E104] (ab32441)
  • Flow Cytometry (Intracellular) - Anti-SIRT1 antibody [E104] (ab32441)
  • Western blot - Anti-SIRT1 antibody [E104] (ab32441)
  • Immunoprecipitation - Anti-SIRT1 antibody [E104] (ab32441)
  • Western blot - Anti-SIRT1 antibody [E104] (ab32441)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIRT1 antibody [E104] (ab32441)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIRT1 antibody [E104] (ab32441)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIRT1 antibody [E104] (ab32441)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIRT1 antibody [E104] (ab32441)
  • Anti-SIRT1 antibody [E104] (ab32441)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [E104] to SIRT1
  • Suitable for: ICC/IF, Flow Cyt (Intra), WB, IP, IHC-P
  • Reacts with: Human

Conjugates logo Related conjugates and formulations

Alexa Fluor® 488 Alexa Fluor® 647 Carrier Free

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Kit
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SIRT1 Activity Assay Kit (Fluorometric) (ab156065)
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Anti-MID1 antibody (ab70770)

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Overview

  • Product name

    Anti-SIRT1 antibody [E104]
    See all SIRT1 primary antibodies
  • Description

    Rabbit monoclonal [E104] to SIRT1
  • Host species

    Rabbit
  • Specificity

    The SIRT1 antibody (ab32441) does not cross-react with other sirtuin family members. Expression levels of the target protein vary with sample type and some optimisation may be required. For western blotting, more concentrated lysates may be required when using tissues samples.

  • Tested applications

    Suitable for: ICC/IF, Flow Cyt (Intra), WB, IP, IHC-Pmore details
  • Species reactivity

    Reacts with: Human
  • Immunogen

    Synthetic peptide within Human SIRT1 aa 700-800 (C terminal). The exact sequence is proprietary.

  • Positive control

    • WB: HeLa, Jurkat, HEK293, SW480, MDA-MB-231 and A549 cell lysates. IHC-P: Human colon carcinoma and human lung squamous carcinoma tissues. Flow: HeLa cells. IP: Jurkat whole cell lysate (ab7899).
  • General notes

    This product is a recombinant monoclonal antibody, which offers several advantages including:

    • - High batch-to-batch consistency and reproducibility
    • - Improved sensitivity and specificity
    • - Long-term security of supply
    • - Animal-free production
    For more information see here.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    Rat: We have preliminary internal testing data to indicate this antibody may not react with this species. Please contact us for more information.

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid freeze / thaw cycles.
  • Storage buffer

    pH: 7.20
    Preservative: 0.01% Sodium azide
    Constituents: 59% PBS, 40% Glycerol, 0.05% BSA
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    E104
  • Isotype

    IgG
  • Research areas

    • Cell Biology
    • Apoptosis
    • Intracellular
    • p53 Pathway
    • Epigenetics and Nuclear Signaling
    • Chromatin Modifying Enzymes
    • Acetylation
    • Microbiology
    • Interspecies Interaction
    • Host Virus Interaction
    • Tags & Cell Markers
    • Subcellular Markers
    • Nucleus
    • Other Nuclear Bodies
    • Epigenetics and Nuclear Signaling
    • Chromatin Modifying Enzymes
    • Acetylation
    • HDACs
    • Class III / Sir2 class
    • Metabolism
    • Types of disease
    • Obesity

Associated products

  • Alternative Versions

    • Alexa Fluor® 647 Anti-SIRT1 antibody [E104] (ab196144)
    • Alexa Fluor® 488 Anti-SIRT1 antibody [E104] (ab196368)
    • Anti-SIRT1 antibody [E104] - BSA and Azide free (ab220807)
  • Assay kits

    • SIRT1 Activity Assay Kit (Fluorometric) (ab156065)
    • Universal SIRT Activity Assay Kit (Colorimetric) (ab156915)
  • Compatible Secondaries

    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab205718)
  • Isotype control

    • Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
  • Positive Controls

    • Jurkat whole cell lysate (ab7899)
  • Recombinant Protein

    • Recombinant human SIRT1 protein (Active) (ab54334)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab32441 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
ICC/IF (1)
1/150.
Flow Cyt (Intra)
1/200.
WB (10)
1/20000. Detects a band of approximately 110 kDa (predicted molecular weight: 82 kDa).

For unpurified, use 1/5000.

Detects a band of approximately 110 kDa (110-121 kDa) which is likely to be due to post translational glycosylation. SIRT1 is known to bind to several other proteins, and the 121kDa band could also be due to the presence of one of these complexes (ensure samples are adequately reduced and denatured).

IP
1/30.
IHC-P (1)
1/150. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Notes
ICC/IF
1/150.
Flow Cyt (Intra)
1/200.
WB
1/20000. Detects a band of approximately 110 kDa (predicted molecular weight: 82 kDa).

For unpurified, use 1/5000.

Detects a band of approximately 110 kDa (110-121 kDa) which is likely to be due to post translational glycosylation. SIRT1 is known to bind to several other proteins, and the 121kDa band could also be due to the presence of one of these complexes (ensure samples are adequately reduced and denatured).

IP
1/30.
IHC-P
1/150. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

Target

  • Function

    NAD-dependent protein deacetylase that links transcriptional regulation directly to intracellular energetics and participates in the coordination of several separated cellular functions such as cell cycle, response to DNA damage, metobolism, apoptosis and autophagy. Can modulate chromatin function through deacetylation of histones and can promote alterations in the methylation of histones and DNA, leading to transcriptional repression. Deacetylates a broad range of transcription factors and coregulators, thereby regulating target gene expression positively and negatively. Serves as a sensor of the cytosolic ratio of NAD(+)/NADH which is altered by glucose deprivation and metabolic changes associated with caloric restriction. Is essential in skeletal muscle cell differentiation and in response to low nutrients mediates the inhibitory effect on skeletal myoblast differentiation which also involves 5'-AMP-activated protein kinase (AMPK) and nicotinamide phosphoribosyltransferase (NAMPT). Component of the eNoSC (energy-dependent nucleolar silencing) complex, a complex that mediates silencing of rDNA in response to intracellular energy status and acts by recruiting histone-modifying enzymes. The eNoSC complex is able to sense the energy status of cell: upon glucose starvation, elevation of NAD(+)/NADP(+) ratio activates SIRT1, leading to histone H3 deacetylation followed by dimethylation of H3 at 'Lys-9' (H3K9me2) by SUV39H1 and the formation of silent chromatin in the rDNA locus. Deacetylates 'Lys-266' of SUV39H1, leading to its activation. Inhibits skeletal muscle differentiation by deacetylating PCAF and MYOD1. Deacetylates H2A and 'Lys-26' of HIST1H1E. Deacetylates 'Lys-16' of histone H4 (in vitro). Involved in NR0B2/SHP corepression function through chromatin remodeling: Recruited to LRH1 target gene promoters by NR0B2/SHP thereby stimulating histone H3 and H4 deacetylation leading to transcriptional repression. Proposed to contribute to genomic integrity via positive regulation of telomere length; however, reports on localization to pericentromeric heterochromatin are conflicting. Proposed to play a role in constitutive heterochromatin (CH) formation and/or maintenance through regulation of the available pool of nuclear SUV39H1. Upon oxidative/metabolic stress decreases SUV39H1 degradation by inhibiting SUV39H1 polyubiquitination by MDM2. This increase in SUV39H1 levels enhances SUV39H1 turnover in CH, which in turn seems to accelerate renewal of the heterochromatin which correlates with greater genomic integrity during stress response. Deacetylates 'Lys-382' of p53/TP53 and impairs its ability to induce transcription-dependent proapoptotic program and modulate cell senescence. Deacetylates TAF1B and thereby represses rDNA transcription by the RNA polymerase I. Deacetylates MYC, promotes the association of MYC with MAX and decreases MYC stability leading to compromised transformational capability. Deacetylates FOXO3 in response to oxidative stress thereby increasing its ability to induce cell cycle arrest and resistance to oxidative stress but inhibiting FOXO3-mediated induction of apoptosis transcriptional activity; also leading to FOXO3 ubiquitination and protesomal degradation. Appears to have a similar effect on MLLT7/FOXO4 in regulation of transcriptional activity and apoptosis. Deacetylates DNMT1; thereby impairs DNMT1 methyltransferase-independent transcription repressor activity, modulates DNMT1 cell cycle regulatory function and DNMT1-mediated gene silencing. Deacetylates RELA/NF-kappa-B p65 thereby inhibiting its transactivating potential and augments apoptosis in response to TNF-alpha. Deacetylates HIF1A, KAT5/TIP60, RB1 and HIC1. Deacetylates FOXO1 resulting in its nuclear retention and enhancement of its transcriptional activity leading to increased gluconeogenesis in liver. Inhibits E2F1 transcriptional activity and apoptotic function, possibly by deacetylation. Involved in HES1- and HEY2-mediated transcriptional repression. In cooperation with MYCN seems to be involved in transcriptional repression of DUSP6/MAPK3 leading to MYCN stabilization by phosphorylation at 'Ser-62'. Deacetylates MEF2D. Required for antagonist-mediated transcription suppression of AR-dependent genes which may be linked to local deacetylation of histone H3. Represses HNF1A-mediated transcription. Required for the repression of ESRRG by CREBZF. Modulates AP-1 transcription factor activity. Deacetylates NR1H3 AND NR1H2 and deacetylation of NR1H3 at 'Lys-434' positively regulates transcription of NR1H3:RXR target genes, promotes NR1H3 proteosomal degradation and results in cholesterol efflux; a promoter clearing mechanism after reach round of transcription is proposed. Involved in lipid metabolism. Implicated in regulation of adipogenesis and fat mobilization in white adipocytes by repression of PPARG which probably involves association with NCOR1 and SMRT/NCOR2. Deacetylates ACSS2 leading to its activation, and HMGCS1. Involved in liver and muscle metabolism. Through deacteylation and activation of PPARGC1A is required to activate fatty acid oxidation in skeletel muscle under low-glucose conditions and is involved in glucose homeostasis. Involved in regulation of PPARA and fatty acid beta-oxidation in liver. Involved in positive regulation of insulin secretion in pancreatic beta cells in response to glucose; the function seems to imply transcriptional repression of UCP2. Proposed to deacetylate IRS2 thereby facilitating its insulin-induced tyrosine phosphorylation. Deacetylates SREBF1 isoform SREBP-1C thereby decreasing its stability and transactivation in lipogenic gene expression. Involved in DNA damage response by repressing genes which are involved in DNA repair, such as XPC and TP73, deacetylating XRCC6/Ku70, and faciliting recruitment of additional factors to sites of damaged DNA, such as SIRT1-deacetylated NBN can recruit ATM to initiate DNA repair and SIRT1-deacetylated XPA interacts with RPA2. Also involved in DNA repair of DNA double-strand breaks by homologous recombination and specifically single-strand annealing independently of XRCC6/Ku70 and NBN. Transcriptional suppression of XPC probably involves an E2F4:RBL2 suppressor complex and protein kinase B (AKT) signaling. Transcriptional suppression of TP73 probably involves E2F4 and PCAF. Deacetylates WRN thereby regulating its helicase and exonuclease activities and regulates WRN nuclear translocation in response to DNA damage. Deacetylates APEX1 at 'Lys-6' and 'Lys-7' and stimulates cellular AP endonuclease activity by promoting the association of APEX1 to XRCC1. Increases p53/TP53-mediated transcription-independent apoptosis by blocking nuclear translocation of cytoplasmic p53/TP53 and probably redirecting it to mitochondria. Deacetylates XRCC6/Ku70 at 'Lys-539' and 'Lys-542' causing it to sequester BAX away from mitochondria thereby inhibiting stress-induced apoptosis. Is involved in autophagy, presumably by deacetylating ATG5, ATG7 and MAP1LC3B/ATG8. Deacetylates AKT1 which leads to enhanced binding of AKT1 and PDK1 to PIP3 and promotes their activation. Proposed to play role in regulation of STK11/LBK1-dependent AMPK signaling pathways implicated in cellular senescence which seems to involve the regulation of the acetylation status of STK11/LBK1. Can deacetylate STK11/LBK1 and thereby increase its activity, cytoplasmic localization and association with STRAD; however, the relevance of such activity in normal cells is unclear. In endothelial cells is shown to inhibit STK11/LBK1 activity and to promote its degradation. Deacetylates SMAD7 at 'Lys-64' and 'Lys-70' thereby promoting its degradation. Deacetylates CIITA and augments its MHC class II transactivation and contributes to its stability. Deacteylates MECOM/EVI1. Deacetylates PML at 'Lys-487' and this deacetylation promotes PML control of PER2 nuclear localization. During the neurogenic transition, repress selective NOTCH1-target genes throug
    Isoform 2: Isoform 2 is shown to deacetylate 'Lys-382' of p53/TP53, however with lower activity than isoform 1. In combination, the two isoforms exert an additive effect. Isoform 2 regulates p53/TP53 expression and cellular stress response and is in turn repressed by p53/TP53 presenting a SIRT1 isoform-dependent auto-regulatory loop.
    (Microbial infection) In case of HIV-1 infection, interacts with and deacetylates the viral Tat protein. The viral Tat protein inhibits SIRT1 deacetylation activity toward RELA/NF-kappa-B p65, thereby potentiates its transcriptional activity and SIRT1 is proposed to contribute to T-cell hyperactivation during infection.
    SirtT1 75 kDa fragment: catalytically inactive 75SirT1 may be involved in regulation of apoptosis. May be involved in protecting chondrocytes from apoptotic death by associating with cytochrome C and interfering with apoptosome assembly.
  • Tissue specificity

    Widely expressed.
  • Sequence similarities

    Belongs to the sirtuin family. Class I subfamily.
    Contains 1 deacetylase sirtuin-type domain.
  • Post-translational
    modifications

    Methylated on multiple lysine residues; methylation is enhanced after DNA damage and is dispensable for deacetylase activity toward p53/TP53.
    Phosphorylated. Phosphorylated by STK4/MST1, resulting in inhibition of SIRT1-mediated p53/TP53 deacetylation. Phosphorylation by MAPK8/JNK1 at Ser-27, Ser-47, and Thr-530 leads to increased nuclear localization and enzymatic activity. Phosphorylation at Thr-530 by DYRK1A and DYRK3 activates deacetylase activity and promotes cell survival. Phosphorylation by mammalian target of rapamycin complex 1 (mTORC1) at Ser-47 inhibits deacetylation activity. Phosphorylated by CaMK2, leading to increased p53/TP53 and NF-kappa-B p65/RELA deacetylation activity (By similarity). Phosphorylation at Ser-27 implicating MAPK9 is linked to protein stability. There is some ambiguity for some phosphosites: Ser-159/Ser-162 and Thr-544/Ser-545.
    Proteolytically cleaved by cathepsin B upon TNF-alpha treatment to yield catalytic inactive but stable SirtT1 75 kDa fragment (75SirT1).
    S-nitrosylated by GAPDH, leading to inhibit the NAD-dependent protein deacetylase activity.
  • Cellular localization

    Cytoplasm. Mitochondrion and Nucleus, PML body. Cytoplasm. Nucleus. Recruited to the nuclear bodies via its interaction with PML (PubMed:12006491). Colocalized with APEX1 in the nucleus (PubMed:19934257). May be found in nucleolus, nuclear euchromatin, heterochromatin and inner membrane (PubMed:15469825). Shuttles between nucleus and cytoplasm (By similarity). Colocalizes in the nucleus with XBP1 isoform 2 (PubMed:20955178).
  • Target information above from: UniProt accession Q96EB6 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 23411 Human
    • Omim: 604479 Human
    • SwissProt: Q96EB6 Human
    • Unigene: 369779 Human
    • Alternative names

      • 75SirT1 antibody
      • hSIR2 antibody
      • hSIRT1 antibody
      • HST2 antibody
      • HST2, S. cerevisiae, homolog of antibody
      • NAD dependent deacetylase sirtuin 1 antibody
      • NAD dependent protein deacetylase sirtuin 1 antibody
      • NAD-dependent deacetylase sirtuin-1 antibody
      • OTTHUMP00000198111 antibody
      • OTTHUMP00000198112 antibody
      • Regulatory protein SIR2 homolog 1 antibody
      • SIR1_HUMAN antibody
      • SIR2 antibody
      • SIR2 like 1 antibody
      • SIR2 like protein 1 antibody
      • SIR2, S.cerevisiae, homolog-like 1 antibody
      • SIR2-like protein 1 antibody
      • SIR2ALPHA antibody
      • SIR2L1 antibody
      • Sirt1 antibody
      • SirtT1 75 kDa fragment antibody
      • Sirtuin (silent mating type information regulation 2 homolog) 1 (S. cerevisiae) antibody
      • Sirtuin 1 antibody
      • Sirtuin type 1 antibody
      see all

    Images

    • Western blot - Anti-SIRT1 antibody [E104] (ab32441)
      Western blot - Anti-SIRT1 antibody [E104] (ab32441)
      All lanes : Anti-SIRT1 antibody [E104] (ab32441) at 1/20000 dilution

      Lane 1 : Wild-type HEK-293 cell lysate
      Lane 2 : SIRT1 CRISPR/Cas9 edited HEK-293 cell lysate
      Lane 3 : MDA-MB-231 cell lysate
      Lane 4 : HeLa cell lysate

      Lysates/proteins at 20 µg per lane.

      Performed under reducing conditions.

      Predicted band size: 82 kDa
      Observed band size: 110 kDa why is the actual band size different from the predicted?



      Lanes 1 - 4: Merged signal (red and green). Green - ab32441 observed at 110 kDa. Red - loading control, ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.

      ab32441 was shown to react with SIRT1 in western blot. The band observed in the CRISPR/Cas9 edited lysate lane below 110kDa may represent truncated forms and cleaved fragments. This has not been investigated further. Membranes were blocked in 3% milk in TBS-T (0.1% Tween®) before incubation with ab32441 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 20000 Dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIRT1 antibody [E104] (ab32441)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIRT1 antibody [E104] (ab32441)

      IHC image of SIRT1 staining in a section of formalin-fixed paraffin-embedded normal human colon* performed on a Leica 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 20mins. The section was then incubated with ab32441, 1/250 dilution, 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 haematoxylin and mounted with DPX.

      For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

      *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

    • Immunocytochemistry/ Immunofluorescence - Anti-SIRT1 antibody [E104] (ab32441)
      Immunocytochemistry/ Immunofluorescence - Anti-SIRT1 antibody [E104] (ab32441)

      Immunofluorescence staining of SH-SY5Y cells with purified ab32441 at a working dilution of 1 in 150, counter-stained with DAPI. The secondary antibody was Alexa Fluor® 488 goat anti rabbit (ab150077), used at a dilution of 1 in 500. The cells were fixed in 4% PFA and permeabilized using 0.1% Triton X 100. The negative control is shown in bottom right hand panel - for the negative control, purified ab32441 was used at a dilution of 1/200 followed by an Alexa Fluor® 594 goat anti-mouse antibody (ab150120) at a dilution of 1/500.

    • Flow Cytometry (Intracellular) - Anti-SIRT1 antibody [E104] (ab32441)
      Flow Cytometry (Intracellular) - Anti-SIRT1 antibody [E104] (ab32441)

      Intracellular Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labeling SIRT1 (red) with ab32441 at a 1/200 dilution. Cells were fixed with 4% paraformaldehyde and permeabilized with 90% methanol. A goat anti-rabbit IgG (Alexa Fluorr® 488) (ab150077) was used as the secondary antibody at a 1/2000 dilution. Black - Rabbit monoclonal IgG (ab172730). Blue (unlabeled control) - Cells without incubation with the primary and secondary antibodies. 

       

       

    • Western blot - Anti-SIRT1 antibody [E104] (ab32441)
      Western blot - Anti-SIRT1 antibody [E104] (ab32441)
      All lanes : Anti-SIRT1 antibody [E104] (ab32441) at 1/20000 dilution (purified)

      Lane 1 : Jurkat cell lysate
      Lane 2 : HeLa cell lysate
      Lane 3 : HEK293 cell lysate
      Lane 4 : A549 cell lysate
      Lane 5 : SW480 cell lysate

      Lysates/proteins at 20 µg per lane.

      Secondary
      All lanes : HRP goat anti-rabbit IgG (H+L) at 1/1000 dilution

      Predicted band size: 82 kDa
      Additional bands at: 110 kDa (possible glycosylated form)



      Blocking buffer and concentration: 5% NFDM/TBST.

      Diluting buffer and concentration: 5% NFDM /TBST.

    • Immunoprecipitation - Anti-SIRT1 antibody [E104] (ab32441)
      Immunoprecipitation - Anti-SIRT1 antibody [E104] (ab32441)

      ab32441 (purified) at 1/30 immunoprecipitating SIRT1 in Jurkat cells (Lane 1). For western blotting, a HRP-conjugated anti-rabbit IgG (H+L) was used as the secondary antibody (1/1000).

      Blocking buffer and concentration: 5% NFDM/TBST.

      Diluting buffer and concentration: 5% NFDM /TBST.

    • Western blot - Anti-SIRT1 antibody [E104] (ab32441)
      Western blot - Anti-SIRT1 antibody [E104] (ab32441)
      All lanes : Anti-SIRT1 antibody [E104] (ab32441) at 1/20000 dilution

      Lane 1 : HeLa whole cell lysate
      Lane 2 : HepG2 whole cell lysate
      Lane 3 : Human Testis tissue lysate
      Lane 4 : Human Colon tissue lysate

      Lysates/proteins at 20 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) preadsorbed at 1/10000 dilution

      Developed using the ECL technique.

      Performed under reducing conditions.

      Predicted band size: 82 kDa
      Observed band size: 110 kDa why is the actual band size different from the predicted?


      Exposure time: 150 seconds


      This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 3% Milk before being incubated with ab32441 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIRT1 antibody [E104] (ab32441)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIRT1 antibody [E104] (ab32441)

      IHC image of SIRT1 staining in a section of formalin-fixed paraffin-embedded normal human colon* performed on a Leica 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 20mins. The section was then incubated with ab32441, 1/250 dilution, 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 haematoxylin and mounted with DPX. The inset secondary-only control image is taken from an identical assay without primary antibody.

      For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

      *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIRT1 antibody [E104] (ab32441)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIRT1 antibody [E104] (ab32441)

      Immunohistochemical staining of paraffin embedded human cerebral cortex with purified ab32441 at a working dilution of 1 in 150. The secondary antibody used is a HRP polymer for rabbit IgG. The sample is counter-stained with hematoxylin. Antigen retrieval was perfomed using Tris-EDTA buffer, pH 9.0. PBS was used instead of the primary antibody as the negative control, and is shown in the inset.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIRT1 antibody [E104] (ab32441)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIRT1 antibody [E104] (ab32441)

      Immunohistochemical analysis of paraffin-embedded human colon carcinoma using unpurified ab32441 at 1/100 dilution.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIRT1 antibody [E104] (ab32441)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-SIRT1 antibody [E104] (ab32441)

      Immunohistochemical analysis of paraffin-embedded human lung squamous carcinoma using unpurified ab32441 at 1/100 dilution.

    • Anti-SIRT1 antibody [E104] (ab32441)
      Anti-SIRT1 antibody [E104] (ab32441)

    Protocols

    To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

    Click here to view the general protocols

    Datasheets and documents

    • SDS download

    • Datasheet download

      Download

    References (126)

    Publishing research using ab32441? Please let us know so that we can cite the reference in this datasheet.

    ab32441 has been referenced in 126 publications.

    • Qazi TJ  et al. Upregulation of mir-132 induces dopaminergic neuronal death via activating SIRT1/P53 pathway. Neurosci Lett 740:135465 (2021). PubMed: 33166640
    • Ye T  et al. Theaflavin protects against oxalate calcium-induced kidney oxidative stress injury via upregulation of SIRT1. Int J Biol Sci 17:1050-1060 (2021). PubMed: 33867828
    • de Carvalho TS  et al. Hypocaloric Diet Initiated Post-Ischemia Provides Long-Term Neuroprotection and Promotes Peri-Infarct Brain Remodeling by Regulating Metabolic and Survival-Promoting Proteins. Mol Neurobiol 58:1491-1503 (2021). PubMed: 33200399
    • Park SA  et al. Role of the SIRT1/p53 regulatory axis in oxidative stress-mediated granulosa cell apoptosis. Mol Med Rep 23:N/A (2021). PubMed: 33179088
    • Tan ML  et al. Autophagy is deregulated in cancer-associated fibroblasts from oral cancer and is stimulated during the induction of fibroblast senescence by TGF-ß1. Sci Rep 11:584 (2021). PubMed: 33436723
    View all Publications for this product

    Customer reviews and Q&As

    Show All Reviews Q&A
    Submit a review Submit a question

    1-10 of 28 Abreviews or Q&A

    Western blot abreview for Anti-SIRT1 antibody [E104]

    Inconclusive
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Mouse Cell lysate - whole cell (CT26 cells)
    Gel Running Conditions
    Reduced Denaturing (4-12% Tris-Bis gel)
    Loading amount
    40 µg
    Specification
    CT26 cells
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C
    Read More

    Abcam user community

    Verified customer

    Submitted Apr 05 2022

    Western blot abreview for Anti-SIRT1 antibody [E104]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Human Cell lysate - whole cell (SHSY5Y cells)
    Gel Running Conditions
    Reduced Denaturing (4-20% tris-glycin gel)
    Loading amount
    40 µg
    Specification
    SHSY5Y cells
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 20°C
    Read More

    Abcam user community

    Verified customer

    Submitted Oct 26 2020

    Western blot abreview for Anti-SIRT1 antibody [E104]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Human Tissue lysate - whole (Placenta)
    Gel Running Conditions
    Reduced Denaturing (10%)
    Loading amount
    20 µg
    Specification
    Placenta
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
    Read More

    Jeremiah Gaudet

    Verified customer

    Submitted Dec 13 2017

    Immunohistochemistry (PFA perfusion fixed frozen sections) abreview for Anti-SIRT1 antibody [E104]

    Poor
    Abreviews
    Abreviews
    abreview image
    Application
    Immunohistochemistry (PFA perfusion fixed frozen sections)
    Sample
    Rat Tissue sections (Brain sections)
    Antigen retrieval step
    None
    Permeabilization
    Yes - 0.3% Triton-X 100
    Specification
    Brain sections
    Blocking step
    (agent) for 45 minute(s) · Concentration: 5% · Temperature: 24°C
    Fixative
    Paraformaldehyde
    Read More

    Vaibhav Konanur

    Verified customer

    Submitted Mar 09 2016

    Western blot abreview for Anti-SIRT1 antibody [E104]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Loading amount
    40 µg
    Gel Running Conditions
    Reduced Denaturing
    Sample
    Rat Tissue lysate - whole (aorta)
    Specification
    aorta
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
    Read More

    Abcam user community

    Verified customer

    Submitted Oct 22 2013

    Western blot abreview for Anti-SIRT1 antibody [E104]

    Excellent
    Abreviews
    Abreviews
    abreview image
    Application
    Western blot
    Sample
    Mouse Cell lysate - whole cell (mouse and human Embryonic stem cell lysate)
    Loading amount
    40 µg
    Specification
    mouse and human Embryonic stem cell lysate
    Gel Running Conditions
    Reduced Denaturing
    Blocking step
    Milk as blocking agent for 3 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
    Read More

    Abcam user community

    Verified customer

    Submitted Jan 03 2013

    Question

    Does your ab32441 react with mouse sample?

    On the sheet it looks not working but today I checked with my mouse ES cell lysate.

    I could detect a band little smaller than human sample. It looks make sense as the protein size.

    Could you check that for me?

    Read More

    Abcam community

    Verified customer

    Asked on Dec 28 2012

    Answer

    Thank you for your inquiry.
    The ab32441 has not been tested in-house on mouse samples. We would not
    predict cross reactivity based on the fact that there is 68% homology
    between our immunogen and the mouse protein, however, we have not determined
    this experimentally. We generally say that anything with > 85% homology
    would be expected to cross-react, but it's not impossible that the antibody
    may bind the mouse protein if enough residues overlap with the antibody.
    I'd recommend that you submit an Abreview with your results to let others
    know about your findings and to earn Abpoints.
    https://www.abcam.com/index.html?pageconfig=resource&rid=10332
    I hope this information helps. Please contact us with any other questions.

    Read More

    Abcam Scientific Support

    Answered on Dec 28 2012

    Question

    Madame, Monsieur,
    Nous avons commandé chez vous 1 Anticorps Anti SIRT1 (ab32441).
    Je n'obtiens aucune bande. J'ai essayé plusieurs concentrations de 25 à 75ug de protéines issues de la culture cellulaire de Fibroblastes humains. Je dilue l'anticorps primaire dans du PBS Tween (0,1%) et du blocking buffer de chez Li-Cor (Volume:Volume). J'ai la aussi essayé plusieurs dilutions différentes de 1/800 à /5000. Je n'ai pas de résultats. Je n'ai bientot plus d'anticorps et n'ai toujours pas de résultats alors que sur les mêmes membranes j'obtiens des bandes pour l'anticorps anti-SIRT4 ab10140.
    Auriez vous des explications à me proposer ? Que puis-je faire pour obtenir un signal ?
    Merci d'avance

    Read More

    Abcam community

    Verified customer

    Asked on Dec 11 2012

    Answer

    Merci de nous avoir contactés et signalé l'anomalie rencontrée avec l'anti-SIRT1 ab32441.

    je pense que cet anticorps aurait du fonctionner en suivant votre protocole et que vous avez malheureusement du recevoir un tube de produit non-fonctionnel.
    Le seul lot actuellement en stock étant celui que vous avez reçu, le GR87015, je peux vous proposer une des solutions suivantes:
    - un remplacement avec une unité de ab110304 : monoclonal de souris, https://www.abcam.com/ab110304
    - un remplacement avec une unité de ab7343 : polyclonal de lapin, https://www.abcam.com/ab7343
    - un avoir ou un remboursement contre la commande originale.

    Merci de m'indiquer la résolution qui vous conviendrait le mieux.

    Read More

    Abcam Scientific Support

    Answered on Dec 11 2012

    Question

    Our technician contacted the ITK diagnostic and they are waiting now for the antibody. Can you please sent it?

    Thanks

    Regards

    Read More

    Abcam community

    Verified customer

    Asked on Aug 24 2012

    Answer

    Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products.

    I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement with the order number 1080353.

    To check the status of the order please contact our Customer Service team and reference this number.

    Please note that this free of charge replacement vial is also covered by our Abpromise guarantee. Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know.

    I wish you the best of luck with your research.

    Read More

    Abcam Scientific Support

    Answered on Aug 24 2012

    Question

    Thanks a lot, I wait the product for the 20th of August.

    Bye

    Read More

    Abcam community

    Verified customer

    Asked on Aug 22 2012

    Answer

    I hope you had good holidays!

    I was placing an agreed order for you however it seems you have purchased this antibody from ITK diagnostics. Could you please let them know Abcam has agreed a free of charge replacement for ab32441 so they should know when it arrives.

    Once you done that please let me know I will validate your order.

    Thank you very much for your cooperation.

    Read More

    Abcam Scientific Support

    Answered on Aug 22 2012

    1-10 of 28 Abreviews or Q&A

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