Anti-ACSS2 antibody (ab66038)
Key features and details
- Rabbit polyclonal to ACSS2
- Suitable for: WB, IHC-P, IP
- Knockout validated
- Reacts with: Human
- Isotype: IgG
Overview
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Product name
Anti-ACSS2 antibody
See all ACSS2 primary antibodies -
Description
Rabbit polyclonal to ACSS2 -
Host species
Rabbit -
Tested Applications & Species
See all applications and species dataApplication Species ICC/IF HumanIHC-P HumanIP HumanWB Human
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Immunogen
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Positive control
- This antibody gave a positive signal in Human Liver and Human Colon Tissue lysates and in the following whole cell lysates: HepG2; U-87 MG; Caco 2.
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General notes
Reproducibility is key to advancing scientific discovery and accelerating scientists’ next breakthrough.
Abcam is leading the way with our range of recombinant antibodies, knockout-validated antibodies and knockout cell lines, all of which support improved reproducibility.
We are also planning to innovate the way in which we present recommended applications and species on our product datasheets, so that only applications & species that have been tested in our own labs, our suppliers or by selected trusted collaborators are covered by our Abpromise™ guarantee.
In preparation for this, we have started to update the applications & species that this product is Abpromise guaranteed for.
We are also updating the applications & species that this product has been “predicted to work with,” however this information is not covered by our Abpromise guarantee.
Applications & species from publications and Abreviews that have not been tested in our own labs or in those of our suppliers are not covered by the Abpromise guarantee.
Please check that this product meets your needs before purchasing. If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, as well as customer reviews and Q&As.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.40
Preservative: 0.02% Sodium azide
Constituent: PBS
Batches of this product that have a concentration
Concentration information loading...Purity
Immunogen affinity purifiedClonality
PolyclonalIsotype
IgGResearch areas
Associated products
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Compatible Secondaries
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Immunizing Peptide (Blocking)
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Isotype control
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab66038 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
GuaranteedTested applications are guaranteed to work and covered by our Abpromise guarantee.
PredictedPredicted to work for this combination of applications and species but not guaranteed.
IncompatibleDoes not work for this combination of applications and species.
Application Species ICC/IF HumanIHC-P HumanIP HumanWB HumanAll applications MouseRatApplication Abreviews Notes WB (1) Use a concentration of 1 µg/ml. Detects a band of approximately 79 kDa (predicted molecular weight: 79 kDa).IHC-P Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.IP (1) Use at an assay dependent concentration.Notes WB
Use a concentration of 1 µg/ml. Detects a band of approximately 79 kDa (predicted molecular weight: 79 kDa).IHC-P
Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.IP
Use at an assay dependent concentration.Application notesIs unsuitable for ICC/IF.Target
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Function
Activates acetate so that it can be used for lipid synthesis or for energy generation. -
Sequence similarities
Belongs to the ATP-dependent AMP-binding enzyme family. -
Cellular localization
Cytoplasm. - Information by UniProt
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Database links
- Entrez Gene: 55902 Human
- Entrez Gene: 60525 Mouse
- Entrez Gene: 311569 Rat
- Omim: 605832 Human
- SwissProt: Q9NR19 Human
- SwissProt: Q8BK97 Mouse
- SwissProt: Q9QXG4 Mouse
- SwissProt: D3ZFF9 Rat
see all -
Alternative names
- ACAS2 antibody
- AceCS antibody
- Acetate CoA ligase antibody
see all
Images
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Western blot - Anti-ACSS2 antibody (ab66038)All lanes : Anti-ACSS2 antibody (ab66038) at 1 µg/ml
Lane 1 : Wild-type HAP1 whole cell lysate
Lane 2 : ACSS2 knockout HAP1 whole cell lysate
Lane 3 : HepG2 whole cell lysate
Lane 4 : U-87 MG whole cell lysate
Lysates/proteins at 20 µg per lane.
Predicted band size: 79 kDa
Observed band size: 80 kDa why is the actual band size different from the predicted?Lanes 1 - 4: Merged signal (red and green). Green - ab66038 observed at 80 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab66038 was shown to recognize ACSS2 in wild-type HAP1 cells as signal was lost at the expected MW in ACSS2 knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and ACSS2 knockout samples were subjected to SDS-PAGE. The membrane was blocked with 3% Milk. Ab66038 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1 ug/ml and 1/20000 dilution respectively. Blots were developed 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 1/20000 dilution for 1 hour at room temperature before imaging.
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Western blot - Anti-ACSS2 antibody (ab66038)All lanes : Anti-ACSS2 antibody (ab66038) at 1 µg/ml
Lane 1 : Human liver tissue lysate - total protein (ab29889)
Lane 2 : HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate
Lane 3 : U-87 MG (Human glioblastoma astrocytoma) Whole Cell Lysate
Lane 4 : Human colon tissue lysate - total protein (ab30051)
Lane 5 :Caco-2 whole cell lysate (ab3950)
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution
Performed under reducing conditions.
Predicted band size: 79 kDa
Observed band size: 79 kDa
Additional bands at: 100 kDa, 38 kDa, 45 kDa. We are unsure as to the identity of these extra bands.
Exposure time: 30 seconds -
Immunoprecipitation - Anti-ACSS2 antibody (ab66038)ACSS2 was immunoprecipitated using 0.5mg HepG2 whole cell extract, 5µg of Rabbit polyclonal to ACSS2 and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).
The antibody was incubated under agitation with Protein G beads for 10min, HepG2 whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.
Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab66038.
Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).
Band: 79kDa: ACSS2. -
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ACSS2 antibody (ab66038)IHC image of ACSS2 staining in human liver carcinoma formalin fixed paraffin embedded tissue section, 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 20 mins. The section was then incubated with ab66038, 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 haematoxylin and mounted with DPX.
Protocols
Datasheets and documents
References (6)
ab66038 has been referenced in 6 publications.
- Wang T et al. SENP1-Sirt3 Signaling Controls Mitochondrial Protein Acetylation and Metabolism. Mol Cell 75:823-834.e5 (2019). PubMed: 31302001
- Kappler M et al. Causes and Consequences of A Glutamine Induced Normoxic HIF1 Activity for the Tumor Metabolism. Int J Mol Sci 20:N/A (2019). PubMed: 31554283
- Sun L et al. Decreased expression of acetyl-CoA synthase 2 promotes metastasis and predicts poor prognosis in hepatocellular carcinoma. Cancer Sci 108:1338-1346 (2017). WB, IHC . PubMed: 28387999
- Chen R et al. Coordinate regulation of stress signaling and epigenetic events by Acss2 and HIF-2 in cancer cells. PLoS One 12:e0190241 (2017). PubMed: 29281714
- Chen R et al. The acetate/ACSS2 switch regulates HIF-2 stress signaling in the tumor cell microenvironment. PLoS One 10:e0116515 (2015). WB . PubMed: 25689462
- Xu M et al. An acetate switch regulates stress erythropoiesis. Nat Med 20:1018-26 (2014). PubMed: 25108527
Images
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Western blot - Anti-ACSS2 antibody (ab66038)All lanes : Anti-ACSS2 antibody (ab66038) at 1 µg/ml
Lane 1 : Wild-type HAP1 whole cell lysate
Lane 2 : ACSS2 knockout HAP1 whole cell lysate
Lane 3 : HepG2 whole cell lysate
Lane 4 : U-87 MG whole cell lysate
Lysates/proteins at 20 µg per lane.
Predicted band size: 79 kDa
Observed band size: 80 kDa why is the actual band size different from the predicted?Lanes 1 - 4: Merged signal (red and green). Green - ab66038 observed at 80 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab66038 was shown to recognize ACSS2 in wild-type HAP1 cells as signal was lost at the expected MW in ACSS2 knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and ACSS2 knockout samples were subjected to SDS-PAGE. The membrane was blocked with 3% Milk. Ab66038 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1 ug/ml and 1/20000 dilution respectively. Blots were developed 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 1/20000 dilution for 1 hour at room temperature before imaging.
-
Western blot - Anti-ACSS2 antibody (ab66038)All lanes : Anti-ACSS2 antibody (ab66038) at 1 µg/ml
Lane 1 : Human liver tissue lysate - total protein (ab29889)
Lane 2 : HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate
Lane 3 : U-87 MG (Human glioblastoma astrocytoma) Whole Cell Lysate
Lane 4 : Human colon tissue lysate - total protein (ab30051)
Lane 5 :Caco-2 whole cell lysate (ab3950)
Lysates/proteins at 10 µg per lane.
Secondary
All lanes : Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution
Performed under reducing conditions.
Predicted band size: 79 kDa
Observed band size: 79 kDa
Additional bands at: 100 kDa, 38 kDa, 45 kDa. We are unsure as to the identity of these extra bands.
Exposure time: 30 seconds -
Immunoprecipitation - Anti-ACSS2 antibody (ab66038)
ACSS2 was immunoprecipitated using 0.5mg HepG2 whole cell extract, 5µg of Rabbit polyclonal to ACSS2 and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).
The antibody was incubated under agitation with Protein G beads for 10min, HepG2 whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.
Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab66038.
Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).
Band: 79kDa: ACSS2. -
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ACSS2 antibody (ab66038)
IHC image of ACSS2 staining in human liver carcinoma formalin fixed paraffin embedded tissue section, 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 20 mins. The section was then incubated with ab66038, 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 haematoxylin and mounted with DPX.
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Immunocytochemistry/ Immunofluorescence - Anti-ACSS2 antibody (ab66038)ICC/IF image of ab66038 stained HeLa cells. The cells were 4% PFA fixed (10 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 (ab66038, 1µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM. This antibody also gave a positive result in 100% methanol fixed (5 min) HeLa cells at 1µg/ml.
