Validated using a knockout cell line
Recombinant
RabMAb

Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)

Abreviews (5) Submit a Question Specific References (25)

Overview

  • Product name
    Anti-APG5L/ATG5 antibody [EPR1755(2)]
    See all APG5L/ATG5 primary antibodies
  • Description
    Rabbit monoclonal [EPR1755(2)] to APG5L/ATG5
  • Host species
    Rabbit
  • Tested applications
    Suitable for: ICC/IF, WB, IP, IHC-Pmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide corresponding to residues in Human APG5L/ATG5.

  • Positive control
    • Raji, HeLa, HT-1080, Human fetal kidney, C6, Raw264.7, PC-12 and NIH3T3 cell lysates; Human hepatocellular carcinoma and Human ovarian adenocarcinoma tissue
  • General notes

    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.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab108327 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 1/100 - 1/250.
WB 1/1000 - 1/10000. Predicted molecular weight: 32 kDa.
IP 1/10 - 1/100.
IHC-P 1/100 - 1/250. Antigen retrieval is recommended.

Target

  • Function
    Involved in autophagic vesicle formation. Conjugation with ATG12, through a ubiquitin-like conjugating system involving ATG7 as an E1-like activating enzyme and ATG10 as an E2-like conjugating enzyme, is essential for its function. The ATG12-ATG5 conjugate acts as an E3-like enzyme which is required for lipidation of ATG8 family proteins and their association to the vesicle membranes. Involved in mitochondrial quality control after oxidative damage, and in subsequent cellular longevity. The ATG12-ATG5 conjugate also negatively regulates the innate antiviral immune response by blocking the type I IFN production pathway through direct association with RARRES3 and MAVS. Also plays a role in translation or delivery of incoming viral RNA to the translation apparatus. Plays a critical role in multiple aspects of lymphocyte development and is essential for both B and T lymphocyte survival and proliferation. Required for optimal processing and presentation of antigens for MHC II. Involved in the maintenance of axon morphology and membrane structures, as well as in normal adipocyte differentiation. Promotes primary ciliogenesis through removal of OFD1 from centriolar satellites and degradation of IFT20 via the autophagic pathway.
    May play an important role in the apoptotic process, possibly within the modified cytoskeleton. Its expression is a relatively late event in the apoptotic process, occurring downstream of caspase activity. Plays a crucial role in IFN-gamma-induced autophagic cell death by interacting with FADD.
  • Tissue specificity
    Ubiquitous. The mRNA is present at similar levels in viable and apoptotic cells, whereas the protein is dramatically highly expressed in apoptotic cells.
  • Sequence similarities
    Belongs to the ATG5 family.
  • Post-translational
    modifications
    Conjugated to ATG12; which is essential for autophagy, but is not required for association with isolation membrane.
    Acetylated by EP300.
  • Cellular localization
    Cytoplasm. Preautophagosomal structure membrane. Colocalizes with nonmuscle actin. The conjugate detaches from the membrane immediately before or after autophagosome formation is completed (By similarity). Localizes also to discrete punctae along the ciliary axoneme and to the base of the ciliary axoneme.
  • Information by UniProt
  • Database links
    see all
  • Alternative names
    • APG 5 antibody
    • APG 5L antibody
    • APG5 antibody
    • APG5 autophagy 5 like antibody
    • APG5 like antibody
    • APG5-like antibody
    • APG5L antibody
    • Apoptosis specific protein antibody
    • Apoptosis-specific protein antibody
    • ASP antibody
    • ATG 5 antibody
    • Atg5 antibody
    • ATG5 autophagy related 5 homolog antibody
    • ATG5_HUMAN antibody
    • Autophagy protein 5 antibody
    • Autophagy related 5 antibody
    • hAPG5 antibody
    • Homolog of S Cerevisiae autophagy 5 antibody
    • OTTHUMP00000040507 antibody
    see all

Images

  • Western blot - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    Western blot - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)

    Lane 1: Wild-type HAP1 cell lysate (20 µg)
    Lane 2: APG5L/ATG5 knockout HAP1 cell lysate (20 µg)
    Lane 3: Raji cell lysate (20 µg)
    Lane 4: Jeg-3 cell lysate (20 µg)

    Lanes 1 - 4: Merged signal (red and green). Green - ab108327 observed at 52 kDa. Red - loading control, ab8245, observed at 37 kDa.
    ab108327 was shown to specifically react with APG5L/ATG5 when APG5L/ATG5 knockout samples were used. Wild-type and APG5L/ATG5 knockout samples were subjected to SDS-PAGE. ab108327 and ab8245 (loading control to GAPDH) were both diluted 1/1000 and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10000 dilution for 1 h at room temperature before imaging.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)

    Unpurified ab108327, at 1/100 dilution, staining APG5L/ATG5 in paraffin-embedded Human ovarian adenocarcinoma tissue by Immunohistochemistry.

  • Immunocytochemistry/ Immunofluorescence - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    Immunocytochemistry/ Immunofluorescence - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)

    Immunofluorescence staining of MCF7 cells with purified ab108327 at a working dilution of 1/150, counter-stained with DAPI. The secondary antibody was Alexa Fluor® 488 goat anti-rabbit (ab150077), used at a dilution of 1/1000. ab7291, a mouse anti-tubulin antibody (1/1000), was used to stain tubulin along with ab150120 (Alexa Fluor® 594 goat anti-mouse, 1/1000), shown in the top right hand panel. The cells were fixed in 100% methanol and permeabilized using 0.1% Triton X 100. The negative controls are shown in bottom middle and right hand panels - for negative control 1, purified ab108327 was used at a dilution of 1/500 followed by an Alexa Fluor® 594 goat anti-mouse antibody (ab150120) at a dilution of 1/500. For negative control 2, ab7291 (mouse anti-tubulin) was used at a dilution of 1/500 followed by an Alexa Fluor® 488 goat anti-rabbit antibody (ab150077) at a dilution of 1/400.

  • Western blot - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    Western blot - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    All lanes : Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327) at 1/10000 dilution (purified)

    Lane 1 : C6 cell lysate
    Lane 2 : PC-12 cell lysate
    Lane 3 : NIH/3T3 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: 32 kDa
    Observed band size: 55 kDa (why is the actual band size different from the predicted?)



    Blocking buffer: 5% NFDM/TBST
    Dilution buffer: 5% NFDM/TBST

  • Western blot - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    Western blot - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327) at 1/10000 dilution (purified) + Raji cell lysate at 20 µg

    Secondary
    HRP goat anti-rabbit IgG (H+L) at 1/10000 dilution

    Predicted band size: 32 kDa
    Observed band size: 55 kDa (why is the actual band size different from the predicted?)



    Blocking buffer: 5% NFDM/TBST
    Dilution buffer: 5% NFDM/TBST

  • Western blot - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    Western blot - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327) at 1/2000 dilution (purified) + HT-1080 cell lysate at 20 µg

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

    Predicted band size: 32 kDa
    Observed band size: 55 kDa (why is the actual band size different from the predicted?)



    Blocking buffer: 5% NFDM/TBST
    Dilution buffer: 5% NFDM/TBST

  • Western blot - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    Western blot - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327) at 1/2000 dilution (purified) + human fetal kidney at 10 µg

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

    Predicted band size: 32 kDa
    Observed band size: 32,55 kDa (why is the actual band size different from the predicted?)



    Blocking buffer: 5% NFDM/TBST
    Dilution buffer: 5% NFDM/TBST

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    Immunohistochemical staining of paraffin embedded human cervical carcinoma with purified ab180327 at a working dilution of 1/150. The secondary antibody used is HRP goat anti-rabbit IgG H&L (ab97051) at 1/500. 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.
  • Immunoprecipitation - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    Immunoprecipitation - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    ab180327 (purified) at 1/20 immunoprecipitating CRSP8 in 10 μg PC-12 whole cell lysate (Lanes 1 and 2, observed at 55 kDa). Lane 3 - PBS. For western blotting, VeriBlot for IP (HRP) (ab131366) was used as the secondary antibody (1/1500). Blocking buffer and concentration: 5% NFDM/TBST Dilution buffer and concentration: 5% NFDM/TBST
  • Western blot - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    Western blot - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    All lanes : Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327) at 1/1000 dilution (unpurified)

    Lane 1 : Raji cell lysate
    Lane 2 : HeLa cell lysate
    Lane 3 : HT-1080 cell lysate
    Lane 4 : Human fetal kidney cell lysate
    Lane 5 : C6 cell lysate
    Lane 6 : Raw264.7 cell lysate
    Lane 7 : PC-12 cell lysate
    Lane 8 : NIH3T3 cell lysate

    Lysates/proteins at 10 µg per lane.

    Predicted band size: 32 kDa



    Secondary antibody - anti-rabbit HRP (ab6721)

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-APG5L/ATG5 antibody [EPR1755(2)] (ab108327)

    Unpurified ab108327, at 1/100 dilution, staining APG5L/ATG5 in paraffin-embedded Human hepatocellular carcinoma tissue by Immunohistochemistry.

References

This product has been referenced in:
  • Liu J  et al. BECN1-dependent CASP2 incomplete autophagy induction by binding to rabies virus phosphoprotein. Autophagy 13:739-753 (2017). Read more (PubMed: 28129024) »
  • Zhang Y  et al. JNK pathway mediates curcumin-induced apoptosis and autophagy in osteosarcoma MG63 cells. Exp Ther Med 14:593-599 (2017). WB . Read more (PubMed: 28672972) »

See all 25 Publications for this product

Publishing research using ab108327? Please let us know so that we can cite the reference in this datasheet.

Customer reviews and Q&As

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Western blot abreview for Anti-APG5L/ATG5 antibody [EPR1755(2)]

 Excellent
Abreviews
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - whole cell (Leukemic Cancer Cell lines)
Gel Running Conditions
Reduced Denaturing (10)
Loading amount
25 µg
Specification
Leukemic Cancer Cell lines
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 23°C
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Submitted Jan 31 2018

Flow Cytometry abreview for Anti-APG5L/ATG5 antibody [EPR1755(2)]

 Good
Abreviews
Application
Flow Cytometry
Sample
Human Cell (Peripheral Blood Mononuclear Cells)
Permeabilization
Yes - 90% Methanol on ice, 20min.
Gating Strategy
FSC v SSC to gate out debris and dying cells followed by CD8+ve selection.
Specification
Peripheral Blood Mononuclear Cells
Preparation
Cell harvesting/tissue preparation method: Cells isolated from whole blood using Dextran/Percoll method.
Sample buffer: 2% FCS
Fixation
Paraformaldehyde
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Submitted Sep 06 2017

Western blot abreview for Anti-APG5L/ATG5 antibody [EPR1755(2)]

 Excellent
Abreviews
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Western blot
Sample
Human Cell lysate - whole cell (Hepato carcinoma HepG2 cells)
Gel Running Conditions
Reduced Denaturing (4 - 15%)
Loading amount
20 µg
Specification
Hepato carcinoma HepG2 cells
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
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Submitted Nov 17 2016

Western blot abreview for Anti-APG5L/ATG5 antibody [EPR1755(2)]

 Good
Abreviews
Application
Western blot
Loading amount
30 µg
Gel Running Conditions
Reduced Denaturing
Sample
Human Cell lysate - whole cell (macrophage)
Specification
macrophage
Treatment
0.1 ug/ml LPS 24 h
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
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Verified customer

Submitted Dec 12 2014

Western blot abreview for Anti-APG5L/ATG5 antibody [EPR1755(2)]

 Average
Abreviews
Application
Western blot
Loading amount
40 µg
Gel Running Conditions
Non-reduced Denaturing (12)
Sample
Xenopus tropicalis Tissue lysate - whole (Brain)
Specification
Brain
Treatment
protease inhibitor cocktail treatment in RIPA lysis buffer
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
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Submitted Mar 24 2014

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