Recombinant Anti-beta III Tubulin antibody [EPR19591] - BSA and Azide free (ab238454)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [EPR19591] to beta III Tubulin - BSA and Azide free
- Suitable for: IHC-P, Flow Cyt (Intra), WB, ICC/IF, IP
- Knockout validated
- Reacts with: Mouse, Rat, Human
Related conjugates and formulations
Overview
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Product name
Anti-beta III Tubulin antibody [EPR19591] - BSA and Azide free
See all beta III Tubulin primary antibodies -
Description
Rabbit monoclonal [EPR19591] to beta III Tubulin - BSA and Azide free -
Host species
Rabbit -
Tested applications
Suitable for: IHC-P, Flow Cyt (Intra), WB, ICC/IF, IPmore details -
Species reactivity
Reacts with: Mouse, Rat, Human -
Immunogen
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- WB: HCT116, HeLa, HEK-293 and HAP1 cell lysates. IHC-P: Human cerebral cortex, glioma, lung adenocarcinoma, and cholangiocarcinoma tissues, mouse cerebral cortex, and rat cerebral cortex tissues. IP: U-87 MG cell lysate. Flow Cyt (intra): U-87 cells, HAP1 cells. ICC/IF: SH-SY5Y and U-87 cells.
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General notes
ab238454 is the carrier-free version of ab215037.
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.
This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.
Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.
This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.
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
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C. Do Not Freeze. -
Storage buffer
pH: 7.2
Constituent: PBS -
Carrier free
Yes -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
EPR19591 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Conjugation kits
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Isotype control
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KO cell lines
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KO cell lysates
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Recombinant Protein
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab238454 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 |
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IHC-P |
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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Flow Cyt (Intra) |
Use at an assay dependent concentration.
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WB |
Use at an assay dependent concentration. Detects a band of approximately 50 kDa (predicted molecular weight: 50 kDa).
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ICC/IF |
Use at an assay dependent concentration.
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IP |
Use at an assay dependent concentration.
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Notes |
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IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol. |
Flow Cyt (Intra)
Use at an assay dependent concentration. |
WB
Use at an assay dependent concentration. Detects a band of approximately 50 kDa (predicted molecular weight: 50 kDa). |
ICC/IF
Use at an assay dependent concentration. |
IP
Use at an assay dependent concentration. |
Target
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Function
Tubulin is the major constituent of microtubules. It binds two moles of GTP, one at an exchangeable site on the beta chain and one at a non-exchangeable site on the alpha-chain. TUBB3 plays a critical role in proper axon guidance and mantainance. -
Tissue specificity
Expression is primarily restricted to central and peripheral nervous system. -
Involvement in disease
Defects in TUBB3 are the cause of congenital fibrosis of extraocular muscles type 3A (CFEOM3A) [MIM:600638]. A congenital ocular motility disorder marked by restrictive ophthalmoplegia affecting extraocular muscles innervated by the oculomotor and/or trochlear nerves. It is clinically characterized by anchoring of the eyes in downward gaze, ptosis, and backward tilt of the head. Congenital fibrosis of extraocular muscles type 3 presents as a non-progressive, autosomal dominant disorder with variable expression. Patients may be bilaterally or unilaterally affected, and their oculo-motility defects range from complete ophthalmoplegia (with the eyes fixed in a hypo- and exotropic position), to mild asymptomatic restrictions of ocular movement. Ptosis, refractive error, amblyopia, and compensatory head positions are associated with the more severe forms of the disorder. In some cases the ocular phenotype is accompanied by additional features including developmental delay, corpus callosum agenesis, basal ganglia dysmorphism, facial weakness, polyneuropathy. -
Sequence similarities
Belongs to the tubulin family. -
Domain
The highly acidic C-terminal region may bind cations such as calcium. -
Post-translational
modificationsSome glutamate residues at the C-terminus are polyglutamylated. This modification occurs exclusively on glutamate residues and results in polyglutamate chains on the gamma-carboxyl group. Also monoglycylated but not polyglycylated due to the absence of functional TTLL10 in human. Monoglycylation is mainly limited to tubulin incorporated into axonemes (cilia and flagella) whereas glutamylation is prevalent in neuronal cells, centrioles, axonemes, and the mitotic spindle. Both modifications can coexist on the same protein on adjacent residues, and lowering glycylation levels increases polyglutamylation, and reciprocally. The precise function of such modifications is still unclear but they regulate the assembly and dynamics of axonemal microtubules. -
Cellular localization
Cytoplasm > cytoskeleton. - Information by UniProt
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Database links
- Entrez Gene: 10381 Human
- Entrez Gene: 22152 Mouse
- Entrez Gene: 246118 Rat
- Omim: 602661 Human
- SwissProt: Q13509 Human
- SwissProt: Q9ERD7 Mouse
- SwissProt: Q4QRB4 Rat
- Unigene: 511743 Human
see all -
Alternative names
- beta 3 tubulin antibody
- beta 4 antibody
- beta-4 antibody
see all
Images
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This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab215037).
Flow cytometry overlay histogram showing wild-type Hap1 (green line) and TUB3 knockout Hap1 stained with ab215037 (red line). The cells were fixed with 4% formaldehyde (10 min) and then permeabilised with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS containing 10% normal goat serum to block non-specific protein-protein interaction followed by the antibody (ab215037) (1x 106 in 100μl at 0.2 μg/ml (1/10400)) for 30min at 22°C.
The secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed was incubated at 1/4000 for 30min at 22°C
Isotype control antibody Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control was used at the same concentration and conditions as the primary antibody (wild-type Hap1 - black line, TUB3 knockout Hap1 - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity).
Acquisition of >5000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter.
This antibody gave a positive signal in Hap1 Fixed with 80% methanol (5 min) / permeabilised with 0.1% PBS-Triton X-100 for 15 min under the same conditions.
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All lanes : Anti-beta III Tubulin antibody [EPR19591] (ab215037) at 1/2000 dilution
Lane 1 : Wild-type HCT116 cell lysate
Lane 2 : TUBB3 knockout HCT116 cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 50 kDa
Observed band size: 52 kDa why is the actual band size different from the predicted?This data was developed using the same antibody clone in a different buffer formulation (ab215037).
Lanes 1- 4: Merged signal (red and green). Green - ab215037 observed at 52 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.
ab215037 was shown to react with beta III Tubulin in wild-type HCT116 cells in western blot. Loss of signal was observed when knockout cell line ab266900 (knockout cell lysate ab257070) was used. Wild-type HCT116 and TUBB3 knockout HCT116 cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab215037 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 2000 dilution and a 1 in 20000 dilution respectively. 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 in 20000 dilution for 1 hour at room temperature before imaging.
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Immunohistochemical analysis of paraffin-embedded human cerebral cortex tissue labeling beta III Tubulin with ab215037 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) at 1/500 dilution.
Cytoplasmic staining on human cerebral cortex is observed.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) at 1/500 dilution.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab215037).
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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All lanes : Anti-beta III Tubulin antibody [EPR19591] (ab215037) at 1/2000 dilution
Lane 1 : Wild-type HeLa cell lysate
Lane 2 : TUBB3 knockout HeLa cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 50 kDa
Observed band size: 50 kDaThis data was developed using the same antibody clone in a different buffer formulation (ab215037).
Lanes 1- 2: Merged signal (red and green). Green - ab215037 observed at 50 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.
ab215037 was shown to react with beta III tubulin in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab255358 (knockout cell lysate ab263857) was used. Wild-type HeLa and TUBB3 knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab215037 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 2000 dilution and a 1 in 20000 dilution respectively. 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 in 20000 dilution for 1 hour at room temperature before imaging.
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All lanes : Anti-beta III Tubulin antibody [EPR19591] (ab215037) at 1/2000 dilution
Lane 1 : Wild-type HAP1 whole cell lysate
Lane 2 : TUBB3 knockout HAP1 whole cell lysate
Lane 3 : HeLa whole cell lysate
Lane 4 : HEK293 whole cell lysate
Lysates/proteins at 20 µg per lane.
Predicted band size: 50 kDaLanes 1 - 4: Merged signal (red and green). Green - ab215037 observed at 50 kDa. Red - loading control, ab9484, observed at 37 kDa.
ab215037 was shown to specifically react with TUBB3 in wild-type HAP1 cells as signal was lost in TUBB3 knockout cells. Wild-type and TUBB3 knockout samples were subjected to SDS-PAGE. ab215037 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at a 1/2000 dilution and 1/20,000 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 1/20,000 dilution for 1 hour at room temperature before imaging.
This data was developed using the same antibody clone in a different buffer formulation containing tissue culture supernatant, tris glycine, BSA, glycerol and sodium azide (ab215037).
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Immunohistochemical analysis of paraffin-embedded human glioma tissue labeling beta III Tubulin with ab215037 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) at 1/500 dilution.
Cytoplasmic staining on human glioma is observed.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) at 1/500 dilution.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab215037).
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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Immunohistochemical analysis of paraffin-embedded human cholangiocarcinoma tissue labeling beta III Tubulin with ab215037 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) at 1/500 dilution.
Cytoplasmic staining on human cholangiocarcinoma is observed.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) at 1/500 dilution.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab215037).
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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Immunohistochemical analysis of paraffin-embedded human hepatocellular carcinoma tissue labeling beta III Tubulin with ab215037 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) at 1/500 dilution.
Negative staining on human hepatocellular carcinoma. (PMID: 25039376).
Counter stained with Hematoxylin.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab215037).
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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Immunohistochemical analysis of paraffin-embedded human lung adenocarcinoma tissue labeling beta III Tubulin with ab215037 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) at 1/500 dilution.
Cytoplasmic staining on human lung adenocarcinoma is observed.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) at 1/500 dilution.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab215037).
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
-
Immunohistochemical analysis of paraffin-embedded mouse cerebral cortex tissue labeling beta III Tubulin with ab215037 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) at 1/500 dilution.
Cytoplasmic staining on mouse cerebral cortex is observed.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) at 1/500 dilution.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab215037).
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
-
Immunohistochemical analysis of paraffin-embedded rat cerebral cortex tissue labeling beta III Tubulin with ab215037 at 1/2000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) at 1/500 dilution.
Cytoplasmic staining on rat cerebral cortex is observed.
Counter stained with Hematoxylin.
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) at 1/500 dilution.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab215037).
Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
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Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized SH-SY5Y (Human neuroblastoma cell line from bone marrow) cells labeling beta III Tubulin with ab215037 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).
Confocal image showing cytoplasmic staining on SH-SY5Y cell line.
The nuclear counterstain is DAPI (blue).
Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594)) at 1/200 dilution (red).
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) at 1/1000 dilution.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab215037).
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Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized U-87 MG (Human glioblastoma-astrocytoma epithelial cell line) cells labeling beta III Tubulin with ab215037 at 1/500 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).
Confocal image showing cytoplasmic staining on U-87 MG cell line.
The nuclear counterstain is DAPI (blue).
Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594)) at 1/200 dilution (red).
Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) at 1/1000 dilution.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab215037).
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Intracellular flow cytometric analysis of 4% paraformaldehyde-fixed U-87 MG (Human glioblastoma-astrocytoma epithelial cell line) cells labeling beta III Tubulin with ab215037 at 1/200 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (Alexa Fluor® 488) at 1/2000 dilution was used as the secondary antibody.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab215037).
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beta III Tubulin was immunoprecipitated from 0.35 mg of U-87 MG (Human glioblastoma-astrocytoma epithelial cell line) whole cell lysate with ab215037 at 1/30 dilution.
Western blot was performed from the immunoprecipitate using ab215037 at 1/1000 dilution.
VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.
Lane 1: U-87 MG whole cell lysate 10µg (Input).
Lane 2: ab215037 IP in U-87 MG whole cell lysate.
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab215037 in U-87 MG whole cell lysate.
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 10 seconds.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab215037).
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.
Datasheets and documents
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Datasheet download
Certificate of Compliance
References (0)
ab238454 has not yet been referenced specifically in any publications.