Western Blot (WB)
(Figure 1. Western blot analysis of Ataxin 3 using anti-Ataxin 3 antibody (PB9423). Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 50ug of sample under reducing conditions. Lane 1: Rat Brain Tissue Lysate Lane 2: COLO320 Whole Cell Lysate Lane 3: HELA Whole Cell Lysate After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA for 50-90 minutes. Blocked the membrane with 5% Non-fat Milk/ TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-Ataxin 3 antigen affinity purified polyclonal antibody at 0.5 ug/mL overnight at 4 degree C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:10000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit with Tanon 5200 system. A specific band was detected for Ataxin 3 at approximately 42KD. The expected band size for Ataxin 3 is at 42KD.)
anti-Human, Rat Ataxin 3 Polyclonal Antibody | anti-ATXN3 antibody
Anti-Ataxin 3 Antibody
Tested Species: Human, Rat
Antigen Retrieval: N/A
Immunohistochemistry - Paraffin: Concentration: 0.5-1ug/ml
Tested Species: Human
Antigen Retrieval: By Heat
Immunohistochemistry - Frozen: Concentration: 0.5-1ug/ml
Tested Species: Human
Antigen Retrieval: N/A
Immunocytochemistry: Concentration: 0.5-1ug/ml
Tested Species: Human
Antigen Retrieval: N/A
Flow Cytometry: 1-3ug/1x106 cells
Tested Species: Human
Antigen Retrieval: N/A
Tested Species: In-house tested species with positive results.
By Heat: Boiling the paraffin sections in 10mM citrate buffer, pH 6.0, for 20 mins is required for the staining of formalin/paraffin sections.
Other applications have not been tested. Optimal dilutions should be determined by end users.
Western Blot (WB)
(Figure 1. Western blot analysis of Ataxin 3 using anti-Ataxin 3 antibody (PB9423). Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 50ug of sample under reducing conditions. Lane 1: Rat Brain Tissue Lysate Lane 2: COLO320 Whole Cell Lysate Lane 3: HELA Whole Cell Lysate After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA for 50-90 minutes. Blocked the membrane with 5% Non-fat Milk/ TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-Ataxin 3 antigen affinity purified polyclonal antibody at 0.5 ug/mL overnight at 4 degree C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:10000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit with Tanon 5200 system. A specific band was detected for Ataxin 3 at approximately 42KD. The expected band size for Ataxin 3 is at 42KD.)
Western Blot (WB)
(Figure 1. Western blot analysis of Ataxin 3 using anti-Ataxin 3 antibody (PB9423). Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 50ug of sample under reducing conditions. Lane 1: Rat Brain Tissue Lysate Lane 2: COLO320 Whole Cell Lysate Lane 3: HELA Whole Cell Lysate After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA for 50-90 minutes. Blocked the membrane with 5% Non-fat Milk/ TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-Ataxin 3 antigen affinity purified polyclonal antibody at 0.5 ug/mL overnight at 4 degree C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:10000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit with Tanon 5200 system. A specific band was detected for Ataxin 3 at approximately 42KD. The expected band size for Ataxin 3 is at 42KD.)
Immunohistochemistry (IHC)
(Figure 2. IHC analysis of Ataxin 3 using anti-Ataxin 3 antibody Ataxin 3 was detected in paraffin-embedded section of Human Lung Cancer Tissue. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1ug/ml rabbit anti-Ataxin 3 Antibody (PB9423) overnight at 4 degree C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37 degree C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) with DAB as the chromogen.)
Immunohistochemistry (IHC)
(Figure 2. IHC analysis of Ataxin 3 using anti-Ataxin 3 antibody Ataxin 3 was detected in paraffin-embedded section of Human Lung Cancer Tissue. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1ug/ml rabbit anti-Ataxin 3 Antibody (PB9423) overnight at 4 degree C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37 degree C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) with DAB as the chromogen.)
Flow Cytometry (FC/FACS)
(Figure 3. Flow Cytometry analysis of A549 cells using anti-ATXN3 antibodyOverlay histogram showing A549 cells stained with PB9423 (Blue line).The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-ATXN3 Antibody (1ug/1x106 cells) for 30 min at 20 degree C. DyLight 488 conjugated goat anti-rabbit IgG (5-10ug/1x106 cells) was used as secondary antibody for 30 minutes at 20 degree C. Isotype control antibody (Green line) was rabbit IgG (1ug/1x106) used under the same conditions. Unlabelled sample (Red line) was also used as a control.)
Flow Cytometry (FC/FACS)
(Figure 3. Flow Cytometry analysis of A549 cells using anti-ATXN3 antibodyOverlay histogram showing A549 cells stained with PB9423 (Blue line).The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-ATXN3 Antibody (1ug/1x106 cells) for 30 min at 20 degree C. DyLight 488 conjugated goat anti-rabbit IgG (5-10ug/1x106 cells) was used as secondary antibody for 30 minutes at 20 degree C. Isotype control antibody (Green line) was rabbit IgG (1ug/1x106) used under the same conditions. Unlabelled sample (Red line) was also used as a control.)
Flow Cytometry (FC/FACS)
(Figure 4. Flow Cytometry analysis of SiHa cells using anti-ATXN3 antibody (PB9423).Overlay histogram showing SiHa cells stained with PB9423 (Blue line).The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-ATXN3 Antibody (1ug/1x106 cells) for 30 min at 20 degree C. DyLight 488 conjugated goat anti-rabbit IgG (5-10ug/1x106 cells) was used as secondary antibody for 30 minutes at 20 degree C. Isotype control antibody (Green line) was rabbit IgG (1ug/1x106) used under the same conditions. Unlabelled sample (Red line) was also used as a control.)
Flow Cytometry (FC/FACS)
(Figure 4. Flow Cytometry analysis of SiHa cells using anti-ATXN3 antibody (PB9423).Overlay histogram showing SiHa cells stained with PB9423 (Blue line).The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-ATXN3 Antibody (1ug/1x106 cells) for 30 min at 20 degree C. DyLight 488 conjugated goat anti-rabbit IgG (5-10ug/1x106 cells) was used as secondary antibody for 30 minutes at 20 degree C. Isotype control antibody (Green line) was rabbit IgG (1ug/1x106) used under the same conditions. Unlabelled sample (Red line) was also used as a control.)
Immunofluorescence (IF)
(Figure 5. IF analysis of ATXN3 using anti- ATXN3 antibody (PB9423)ATXN3 was detected in immunocytochemical section of U20S cell. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent for 15 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2ug/mL rabbit anti- ATXN3 Antibody overnight at 4 degree C. DyLight 488 Conjugated Goat Anti-Rabbit IgG was used as secondary antibody at 1:100 dilution and incubated for 30 minutes at 37 degree C. Visualize using a fluorescence microscope and filter sets appropriate for the label used.)
Immunofluorescence (IF)
(Figure 5. IF analysis of ATXN3 using anti- ATXN3 antibody (PB9423)ATXN3 was detected in immunocytochemical section of U20S cell. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent for 15 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2ug/mL rabbit anti- ATXN3 Antibody overnight at 4 degree C. DyLight 488 Conjugated Goat Anti-Rabbit IgG was used as secondary antibody at 1:100 dilution and incubated for 30 minutes at 37 degree C. Visualize using a fluorescence microscope and filter sets appropriate for the label used.)
Background: ATXN3 (Ataxin 3), also known as AT3, MJD GENE, MJD1, SCA3 GENE, ATX3, JOS, Spinocerebellar ataxia-3, Machado-Joseph disease protein 1, is a protein that in humans is encoded by the ATXN3 gene. ATXN3 ranges in size from 360 to 374 amino acids. Using Northern blot analysis showed that ATXN3 mRNA was ubiquitously expressed in human tissues. They detected at least 4 ATXN3 transcripts of 1.4, 1.8, 4.5, and 7.5 kb and suggested that the different mRNA species probably result from differential splicing and polyadenylation. Machado-Joseph disease, also known as spinocerebellar ataxia-3, is an autosomal dominant neurologic disorder. The protein encoded by the ATXN3 gene contains (CAG)n repeats in the coding region, and the expansion of these repeats from the normal 13-36 to 68-79 is the cause of Machado-Joseph disease. There is an inverse correlation between the age of onset and CAG repeat numbers. Alternatively spliced transcript variants encoding different isoforms have been described for this gene. Ataxin-3 interacted with 2 human homologs of the yeast DNA repair protein RAD23, HHR23A (RAD23A) and HHR23B (RAD23B). Both normal and mutant ataxin-3 proteins interacted with the ubiquitin-like domain at the N terminus of the HHR23 proteins, which is a motif important for nucleotide excision repair. However, in HEK 293 cells, HHR23A was recruited to intranuclear inclusions formed by the mutant ataxin-3 through its interaction with ataxin-3.
NCBI and Uniprot Product Information
NCBI Description
Machado-Joseph disease, also known as spinocerebellar ataxia-3, is an autosomal dominant neurologic disorder. The protein encoded by this gene contains (CAG)n repeats in the coding region, and the expansion of these repeats from the normal 13-36 to 68-79 is one cause of Machado-Joseph disease. There is a negative correlation between the age of onset and CAG repeat numbers. Alternatively spliced transcript variants encoding different isoforms have been described for this gene. [provided by RefSeq, Sep 2009]
Uniprot Description
ataxin-3: Deubiquitinating enzyme involved in protein homeostasis maintenance, transcription, cytoskeleton regulation, myogenesis and degradation of misfolded chaperone substrates. Binds long polyubiquitin chains and trims them, while it has weak or no activity against chains of 4 or less ubiquitins. Involved in degradation of misfolded chaperone substrates via its interaction with STUB1/CHIP: recruited to monoubiquitinated STUB1/CHIP, and restricts the length of ubiquitin chain attached to STUB1/CHIP substrates and preventing further chain extension. In response to misfolded substrate ubiquitination, mediates deubiquitination of monoubiquitinated STUB1/CHIP. Interacts with key regulators of transcription and represses transcription: acts as a histone- binding protein that regulates transcription. Defects in ATXN3 are the cause of spinocerebellar ataxia type 3 (SCA3); also known as Machado-Joseph disease (MJD). Spinocerebellar ataxia is a clinically and genetically heterogeneous group of cerebellar disorders. Patients show progressive incoordination of gait and often poor coordination of hands, speech and eye movements, due to degeneration of the cerebellum with variable involvement of the brainstem and spinal cord. SCA3 belongs to the autosomal dominant cerebellar ataxias type I (ADCA I) which are characterized by cerebellar ataxia in combination with additional clinical features like optic atrophy, ophthalmoplegia, bulbar and extrapyramidal signs, peripheral neuropathy and dementia. The molecular defect in SCA3 is the a CAG repeat expansion in ATXN3 coding region. Longer expansions result in earlier onset and more severe clinical manifestations of the disease. 3 isoforms of the human protein are produced by alternative splicing.
Protein type: Ubiquitin-specific protease; DNA repair, damage; Transcription regulation; Protease; EC 3.4.19.12
Chromosomal Location of Human Ortholog: 14q21
Cellular Component: cytoplasm; cytosol; endoplasmic reticulum membrane; mitochondrial matrix; mitochondrial membrane; nuclear inclusion body; nuclear matrix; nucleoplasm; nucleus
Molecular Function: ATPase binding; histone deacetylase activity; identical protein binding; omega peptidase activity; protein binding; ubiquitin protein ligase binding; ubiquitin-specific protease activity
Biological Process: actin cytoskeleton organization and biogenesis; intermediate filament cytoskeleton organization and biogenesis; microtubule cytoskeleton organization and biogenesis; misfolded or incompletely synthesized protein catabolic process; nervous system development; nucleotide-excision repair; proteasomal ubiquitin-dependent protein catabolic process; protein deubiquitination; regulation of transcription, DNA-dependent; synaptic transmission; transcription, DNA-dependent; ubiquitin-dependent protein catabolic process
Disease: Machado-joseph Disease
Research Articles on ATXN3
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Product Notes
The ATXN3 atxn3 (Catalog #AAA178269) is an Antibody and is intended for research purposes only. The product is available for immediate purchase. The Anti-Ataxin 3 Antibody reacts with Human, Rat and may cross-react with other species as described in the data sheet. AAA Biotech's Ataxin 3 can be used in a range of immunoassay formats including, but not limited to, Western Blot (WB), Immunohistochemistry (IHC) Paraffin/Frozen, Immunocytochemistry (ICC), Flow Cytometry (FC). Western Blot: Concentration: 0.1-0.5ug/ml Tested Species: Human, Rat Antigen Retrieval: N/A Immunohistochemistry - Paraffin: Concentration: 0.5-1ug/ml Tested Species: Human Antigen Retrieval: By Heat Immunohistochemistry - Frozen: Concentration: 0.5-1ug/ml Tested Species: Human Antigen Retrieval: N/A Immunocytochemistry: Concentration: 0.5-1ug/ml Tested Species: Human Antigen Retrieval: N/A Flow Cytometry: 1-3ug/1x106 cells Tested Species: Human Antigen Retrieval: N/A Tested Species: In-house tested species with positive results. By Heat: Boiling the paraffin sections in 10mM citrate buffer, pH 6.0, for 20 mins is required for the staining of formalin/paraffin sections. Other applications have not been tested. Optimal dilutions should be determined by end users. Researchers should empirically determine the suitability of the ATXN3 atxn3 for an application not listed in the data sheet. Researchers commonly develop new applications and it is an integral, important part of the investigative research process. It is sometimes possible for the material contained within the vial of "Ataxin 3, Polyclonal Antibody" to become dispersed throughout the inside of the vial, particularly around the seal of said vial, during shipment and storage. We always suggest centrifuging these vials to consolidate all of the liquid away from the lid and to the bottom of the vial prior to opening. Please be advised that certain products may require dry ice for shipping and that, if this is the case, an additional dry ice fee may also be required.Precautions
All products in the AAA Biotech catalog are strictly for research-use only, and are absolutely not suitable for use in any sort of medical, therapeutic, prophylactic, in-vivo, or diagnostic capacity. By purchasing a product from AAA Biotech, you are explicitly certifying that said products will be properly tested and used in line with industry standard. AAA Biotech and its authorized distribution partners reserve the right to refuse to fulfill any order if we have any indication that a purchaser may be intending to use a product outside of our accepted criteria.Disclaimer
Though we do strive to guarantee the information represented in this datasheet, AAA Biotech cannot be held responsible for any oversights or imprecisions. AAA Biotech reserves the right to adjust any aspect of this datasheet at any time and without notice. It is the responsibility of the customer to inform AAA Biotech of any product performance issues observed or experienced within 30 days of receipt of said product. To see additional details on this or any of our other policies, please see our Terms & Conditions page.Item has been added to Shopping Cart
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