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Anti-beta Actin antibody (ab8227)
Product name
Anti-beta Actin antibody
Description
Rabbit polyclonal to beta Actin
Specificity
The immunogen used for this product shares 77% homology with Gamma actin/actin cytoplasmic 2. Cross-reactivity with this protein has not been confirmed experimentally.
Tested applications
Suitable for:
IHC-Fr, IP, WB, ICC, Flow Cyt, IHC-FrFl, IHC-P, IHC-P, ICC/IF, ELISA
Species reactivity
Reacts with:
Mouse, Rat, Sheep, Rabbit, Chicken, Guinea pig, Cow, Dog, Human, Pig, Xenopus laevis, Drosophila melanogaster, Fish, Monkey, Zebrafish, Rhesus monkey, Chinese hamster
Synthetic peptide (the amino acid sequence is considered to be commercially sensitive) within Human beta Actin aa 1-100. The exact sequence is proprietary.(Peptide available as , )
Positive control
WB: Mouse brain tissue lysate and HeLa, A431, HEK293, NIH3T3 and PC12 whole cell lysates.
IHC-P: Normal human colon and normal human liver tissues.
ICC/IF: NIH3T3 and SV40LT-SMC cells.
Properties
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
Preservative: 0.02% Sodium Azide
Constituents: 1% BSA, PBS, pH 7.4
Immunogen affinity purified
Polyclonal
Research areas
Associated products
Alternative Products
Compatible Secondaries
Immunizing Peptide (Blocking)
Related Products
Applications
covers the use of
in the following tested applications.
The application notes include recommende optimal dilutions/concentrations should be determined by the end user.
Application
Use at an assay dependent concentration.
Use at an assay dependent concentration.
1/1000 - 1/5000.
Use at an assay dependent concentration.
Use at an assay dependent concentration.
- Rabbit polyclonal IgG, is suitable for use as an isotype control with this antibody.
Use at an assay dependent concentration.
Use a concentration of 1 ug/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
Use a concentration of 0.2 ug/ml.
Use a concentration of 1 ug/ml.
Actins are highly conserved proteins that are involved in various types of cell motility and are ubiquitously expressed in all eukaryotic cells.
Involvement in disease
Defects in ACTB are a cause of dystonia juvenile-onset (DYTJ) [MIM:607371]. DYTJ is a form of dystonia with juvenile onset. Dystonia is defined by the presence of sustained involuntary muscle contraction, often leading to abnormal postures. DYTJ patients manifest progressive, generalized, dopa-unresponsive dystonia, developmental malformations and sensory hearing loss.
Sequence similarities
Belongs to the actin family.
Post-translationalmodifications
ISGylated.
Cellular localization
Cytoplasm & cytoskeleton. Localized in cytoplasmic mRNP granules containing untranslated mRNAs.
Database links
Alternative names
A26C1A antibody
A26C1B antibody
ACTB antibody
Anti-beta Actin antibody images
Western blot - Anti-beta Actin antibody (ab8227)
All lanes : Anti-beta Actin antibody (ab8227) at 1/1000 dilutionLane 1 : A431 (Human epithelial carcinoma cell line) Whole Cell LysateLane 2 : HEK293 (Human embryonic kidney cell line) Whole Cell LysateLane 3 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell LysateLane 4 : PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell LysateLysates/proteins at 20 ug per lane.SecondaryGoat Anti-Rabbit IgG H&L (Alexa Fluor(R) 790) () at 1/10000 dilution
Immunocytochemistry/ Immunofluorescence - Anti-beta Actin antibody (ab8227)
ab8227 staining beta Actin in SV40LT-SMC cells. The cells were fixed with 4% formaldehyde (10min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at +4&C with ab8227 at 1&g/ml (shown in green) and , Mouse monoclonal [DM1A] to alpha Tubulin - Microtubule Marker (Alexa Fluor& 594), at 1/250 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-beta Actin antibody (ab8227)
IHC image of ab8227 staining beta Actin in rat small intestine formalin fixed paraffin embedded tissue sections, performed on a Leica Bond. The section was pre-treated using heat mediated antigen retrieval with EDTA (epitope retrieval solution 2) for 20 mins. The section was then incubated with ab&g/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. No primary antibody was used in the secondary only control (shown on the inset).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.
Western blot - Anti-beta Actin antibody (ab8227)
All lanes : Anti-beta Actin antibody (ab8227) at 0.1 ug/mlLane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell LysateLane 2 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell LysateLane 3 : Liver (Rat) Tissue LysateLysates/proteins at 10 ug per lane.SecondaryGoat Anti-Rabbit IgG H&L (HRP) () at 1/50000 dilutionPerformed under reducing conditions.Exposure time :
Immunocytochemistry/ Immunofluorescence - Anti-beta Actin antibody (ab8227)
ab8227 staining beta Actin in NIH3T3 cells. The cells were fixed with 100% methanol (5min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at +4&C with ab8227 at 1&g/ml (shown in green) and , Mouse monoclonal [DM1A] to alpha Tubulin - Microtubule Marker (Alexa Fluor& 594), at 1/250 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-beta Actin antibody (ab8227)
IHC image of beta actin staining in a section of formalin-fixed paraffin-embedded&normal human&colon*. The section was pre-treated using pressure cooker heat mediated antigen retrieval with sodium citrate buffer (pH6) for 30mins. The section was then incubated with ab0 dilution, for 15 mins at room temperature. A goat anti-rabbit biotinylated secondary antibody (, 1/1000 dilution) was used to detect the primary, and visualized using an HRP conjugated ABC system. Streptavidin HRP was used,
at a 1/10000 dilution. DAB was used as the chromogen (), diluted 1/100 and incubated for 10min at room temperature. The section was then counterstained with haematoxylin and mounted with DPX.The inset negative 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
Western blot - Anti-beta Actin antibody (ab8227)
All lanes : Anti-beta Actin antibody (ab8227) at 1/1000 dilutionLane 1 : HeLa nuclear lysateLane 2 : HeLa whole cell lysateLane 3 : A431 cell lysateLane 4 : Jurkat cell lysateLane 5 : HEK 293 cell lysateLane 6 : HeLa nuclear lysate with Human beta Actin peptide () at 1 ug/mlLane 7 : HeLa whole cell lysate with Human beta Actin peptide () at 1 ug/mlLane 8 : A431 cell lysate with Human beta Actin peptide () at 1 ug/mlLane 9 : Jurkate cell lysate with Human beta Actin peptide () at 1 ug/mlLane 10 : HEK 293 cell lysate with Human beta Actin peptide () at 1 ug/mlLysates/proteins at 20 ug per lane.SecondaryGoat Anti-Rabbit IgG H&L (HRP) () at 1/5000 dilutionExposure time :
Immunocytochemistry/ Immunofluorescence - Anti-beta Actin antibody (ab8227)
ICC/IF image of ab8227 stained human HeLa cells. The cells were methanol fixed (5 min) and incubated with the antibody (ab8227, 1&g/ml) for 1h at room temperature. The secondary antibody (green)&was Alexa Fluor& 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Image-iTTM FX Signal Enhancer was used as the primary blocking agent, 5% BSA was used for all other blocking steps. DAPI was used to stain the cell nuclei (blue).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-beta Actin antibody (ab8227)
IHC image of beta Actin staining in normal human colon, formalin-fixed and paraffin-embedded tissue*. The section was pre-treated using pressure cooker heat mediated antigen retrieval with sodium citrate buffer (pH6) for 30mins. The section was incubated with ab8227, 3&g/ml overnight at +4&C. A secondary antibody&(Ab9 dilution) was used for 1hr at room temperature. The section was counterstained with haematoxylin and mounted with DPX.The inset negative control image is taken from an identical assay without primary antibody.*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre
Western blot - Anti-beta Actin antibody (ab8227)
All lanes : Anti-beta Actin antibody (ab8227) at 1/1000 dilutionLane 1 : HeLa nuclear lysate at 20 &g/mlLane 2 : HeLa whole cell lysate at 20 &g/mlLane 3 : A431 cell lysate at 20 &g/mlLane 4 : Jurkat cell lysate at 20 &g/mlLane 5 : HEK 293 cell lysate at 20 &g/mlLane 6 : HeLa nuclear lysate at 20 &g/ml with Human beta Actin peptide () at 1 &g/mlLane 7 : HeLa whole cell lysate at 20 &g/ml with Human beta Actin peptide () at 1 &g/mlLane 8 : A431 cell lysate at 20 &g/ml with Human beta Actin peptide () at 1 &g/mlLane 9 : Jurkate cell lysate at 20 &g/ml with Human beta Actin peptide () at 1 &g/mlLane 10 : HEK 293 cell lysate at 20 &g/ml with Human beta Actin peptide () at 1 &g/mlSecondaryGoat Anti-Rabbit IgG H&L (HRP) () at 1/5000 dilution
Western blot - Anti-beta Actin antibody (ab8227)
All lanes : Anti-beta Actin antibody (ab8227) at 1/1000 dilutionLane 1 : HeLa cells (Human) at 20 ugLane 2 : 3T3 cells (Mouse) at 20 ugLane 3 : Fish Liver at 20 ugLane 4 : Rabbit Liver at 20 ugLane 5 : MDCK cells (Dog) at 20 ug/mlLane 6 : EBTr cells (Cow) at 20 ugLane 7 : SL-29 cells (Chicken) at 20 ug/mlLane 8 : CHO cells (Chinese Hamster) at 20 ugLane 9 : Xenopus embryo at 20 ugSecondaryGoat Anti-Rabbit IgG H&L (HRP) () at 1/5000 dilutionExposure time :
30 seconds
Immunohistochemistry (Frozen sections) - Anti-beta Actin antibody (ab8227)This image is courtesy of an anonymous abreview.
ab8227 staining beta Actin in human stomach tissue sections by Immunohistochemistry (frozen sections). Tissue was fixed with acetone and then blocked with 5% serum for 1 hour at 23&C followed by incubation with the primary antibody, at 1&g/ml, for 1 hour at 23&C. A diluted anti-rabbit HRP-conjugated goat polyclonal was used as secondary antibody.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-beta Actin antibody (ab8227)
IHC image of beta Actin staining in human liver 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 ab8227, 1ug/ml, for 8 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.
ELISA - Anti-beta Actin antibody (ab8227)Image courtesy of an anonymous Abreview.
ab8227 used in Direct ELISA in NIH 3T3 murine fibroblasts. Primary antibody used at a 1/1000 dilution for 16 hours at 4&C. The secondary antibody used is an AP-conjugated Goat anti-rabbit used at a 1/2000 dilution. A blocking step was performed using 5% BSA for 1 hour at 23&C.
References for Anti-beta Actin antibody (ab8227)
This product has been referenced in:
Sun Y &et al. Therapeutic effect of apocynin through antioxidant activity and suppression of apoptosis and inflammation after spinal cord injury. Exp Ther Med 13:952-960 (2017).
Kostarnoy AV &et al. Receptor Mincle promotes skin allergies and is capable of recognizing cholesterol sulfate. Proc Natl Acad Sci U S A 114:E (2017).
99 Abreviews or Q&A
Application
Western blot
Loading amount
Gel Running Conditions
Reduced Denaturing
Zebrafish Tissue lysate - whole (Larvae 5hpf)
Specification
Larvae 5hpf
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) & Concentration: 5% & Temperature: 21&C
Mr. Pedro Guedes Dias
Verified customer
Application
Western blot
Loading amount
Gel Running Conditions
Reduced Denaturing
Rat Cell lysate - whole cell (Primary Neurons)
Specification
Primary Neurons
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) & Concentration: 5% & Temperature: 21&C
Abcam user community
Verified customer
Application
Immunohistochemistry (Frozen sections)
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) & Concentration: 5% & Temperature: 23&C
Mouse Tissue sections (Skin)
Specification
Permeabilization
Yes - 0,3% TritonX
Perfusion fixed with paraformaldehyde and picric acid
Mr. Frank Rommel
Verified customer
Application
Western blot
Loading amount
Gel Running Conditions
Reduced Denaturing
(Biorad Criterion TGX Stain Free Gel, Any kD, Cat. number 5678123)
Fruit fly (Drosophila melanogaster) Tissue lysate - whole (Drosophila heads)
Specification
Drosophila heads
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) & Concentration: 5% & Temperature: 20&C
Abcam user community
Verified customer
this product to work in the species/application used in this Abreview.
Application
Western blot
Loading amount
Gel Running Conditions
Reduced Denaturing
Rat Tissue lysate - whole (ovaries)
Specification
Blocking step
Milk as blocking agent for 18 hour(s) and 0 minute(s) & Concentration: 10% & Temperature: 4&C
Mrs. Henda Nabli
Verified customer
Abcam community
Verified customer
We do not know if ab8227 stains nuclear actin. It is raised against a sequence from cytoplasmic actin, and we have not tested for cross-reactivity with nuclear actin.
http://www.uniprot.org/uniprot/P60709
Abcam Scientific Support
Abcam community
Verified customer
I just received an update form lab regarding the epitope for ab79667 and ab113279. Unfortunately the epitope has not been determined so the information is not available. I am sorry we will not be able to suggest these to be paired with ab8227 in sELISA.
Abcam Scientific Support
Abcam community
Verified customer
I have had the opportunity to discuss our QC methods regarding this product with the lab. Ab17721 is batch tested by us in WB in HeLa, Jurkat and NIH3T3 lysates and also in Immunofluorescence in HeLa cells. Each new AP purified must pass both of these ...
Abcam Scientific Support
Abcam has not validated the combination of species/application used in this Abreview.
Application
Western blot
Bat Cell lysate - whole cell (kidney)
Gel Running Conditions
Reduced Denaturing
(10% SDS-PAGE)
Loading amount
20000 cells
Specification
Blocking step
BSA as blocking agent for 18 hour(s) and 0 minute(s) & Concentration: 5% & Temperature: 4&C
Noreen Rapin
Verified customer
Application
Western blot
Human Cell lysate - whole cell (HEK293T)
Gel Running Conditions
Reduced Denaturing
Loading amount
Specification
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) & Concentration: 5% & Temperature: 20&C
Abcam user community
Verified customer
1-10 of 99 Abreviews or Q&A
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Please note: All products are &FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE&
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