Ubiquitin Recombinant Rabbit Monoclonal Antibody [ST46-03] - BSA and Azide free
Catalog# HA750165
Ubiquitin Recombinant Rabbit Monoclonal Antibody [ST46-03] - BSA and Azide free
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WB
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IF-Cell
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IF-Tissue
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IHC-P
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FC
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Human
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Mouse
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Rat
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ET1609-21
含抗保成分
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unconjugated
Overview
Product Name
Ubiquitin Recombinant Rabbit Monoclonal Antibody [ST46-03] - BSA and Azide free
Antibody Type
Recombinant Rabbit monoclonal Antibody
Immunogen
Synthetic peptide within human Ubiquitin aa 1-50.
Species Reactivity
Human, Mouse, Rat
Validated Applications
WB, IF-Cell, IF-Tissue, IHC-P, FC
Positive Control
HeLa cell lysate, HeLa treated with 10μM MG-132 for 6 hours cell lysate, NIH/3T3 cell lysate, NIH/3T3 treated with 10μM MG-132 for 8 hours cell lysate, PC-12 cell lysate, 293T cell lysate, HepG2 cell lysate, HepG2, NIH/3T3 cells treated with 10µM MG-132 for 6 hours, PC-12 cells treated with 10µM MG-132 for 6 hours, human esophagus tissue, mouse esophagus tissue, rat esophagus tissue, mouse brain tissue, rat brain tissue.
Conjugation
unconjugated
Clone Number
ST46-03
Product Features
Form
Liquid
Concentration
1mg/ml
Storage Instructions
Store at +4℃ after thawing. Aliquot store at -20℃ or -80℃. Avoid repeated freeze / thaw cycles.
Storage Buffer
1*PBS (pH7.4).
Isotype
IgG
Purification Method
Protein A affinity purified.
Application Dilution
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WB
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1:5,000-1:10,000
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IF-Cell
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1:200-1:500
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IF-Tissue
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1:500-1:2,000
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IHC-P
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1:1,0000
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FC
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1:1,000
Target
Function
Ubiquitin is a conserved polypeptide unit that plays an important role in the ubiquitin-proteasome pathway. Ubiquitin exists either covalently attached to another protein, or free (unanchored). When covalently bound, it is conjugated to target proteins via an isopeptide bond either as a monomer (monoubiquitin), a polymer linked via different Lys residues of the ubiquitin (polyubiquitin chains) or a linear polymer linked via the initiator Met of the ubiquitin (linear polyubiquitin chains). Polyubiquitin chains, when attached to a target protein, have different functions depending on the Lys residue of the ubiquitin that is linked: Lys-6-linked may be involved in DNA repair; Lys-11-linked is involved in ERAD (endoplasmic reticulum-associated degradation) and in cell-cycle regulation; Lys-29-linked is involved in lysosomal degradation; Lys-33-linked is involved in kinase modification; Lys-48-linked is involved in protein degradation via the proteasome; Lys-63-linked is involved in endocytosis, DNA-damage responses as well as in signaling processes leading to activation of the transcription factor NF-kappa-B. This antibody reacts with ubiquitin, a polypeptide w/ Mr. of approx. 8.5kD. It reacts with physiologically occurring conjugates of ubiquitin and intracellular proteins. Specifically recognizes ubiquitinated cytoplasmic inclusion bodies.
Background References
1. Zhang P et al. Endometrial cancer-associated mutants of SPOP are defective in regulating estrogen receptor-a protein turnover. Cell Death Dis 6:e1687 (2015).
2. Ye X et al. Characterization of PHB1 and its role in mitochondrial maturation and yolk platelet degradation during development of Artemia embryos. PLoS One 9:e109152 (2014).
Sequence Similarity
Belongs to the ubiquitin family.
Post-translational Modification
[Ubiquitin]: Phosphorylated at Ser-65 by PINK1 during mitophagy. Phosphorylated ubiquitin specifically binds and activates parkin (PRKN), triggering mitophagy. Phosphorylation does not affect E1-mediated E2 charging of ubiquitin but affects discharging of E2 enzymes to form polyubiquitin chains. It also affects deubiquitination by deubiquitinase enzymes such as USP30.; [Ubiquitin]: Mono-ADP-ribosylated at the C-terminus by PARP9, a component of the PPAR9-DTX3L complex. ADP-ribosylation requires processing by E1 and E2 enzymes and prevents ubiquitin conjugation to substrates such as histones.
Subcellular Location
Cytoplasm, Nucleus, Membrane, Mitochondrion.
UNIPROT
Synonyms
FLJ25987 antibody
MGC8385 antibody
Polyubiquitin B antibody
RPS 27A antibody
RPS27A antibody
UBA 52 antibody
UBA 80 antibody
UBA52 antibody
UBA80 antibody
UBB antibody
ExpandImages
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☑ Cell treatment (CT)
Western blot analysis of Ubiquitin on different lysates with Rabbit anti-Ubiquitin antibody (HA750165) at 1/5,000 dilution and competitor's antibody at 1/2,000 dilution.
Lane 1: HeLa cell lysate
Lane 2: HeLa treated with 10μM MG-132 for 6 hours cell lysate
Lane 3: NIH/3T3 cell lysate
Lane 4: NIH/3T3 treated with 10μM MG-132 for 8 hours cell lysate
Lane 5: PC-12 cell lysate
Lysates/proteins at 20 µg/Lane.
Exposure time: 1 minute 34 seconds; ECL: K1801;
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA750165) at 1/5,000 dilution and competitor's antibody at 1/2,000 dilution were used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature. -
Immunocytochemistry analysis of HepG2 cells labeling Ubiquitin with Rabbit anti-Ubiquitin antibody (HA750165) at 1/500 dilution.
Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Ubiquitin antibody (HA750165) at 1/500 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.
Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution. -
☑ Cell treatment (CT)
Immunocytochemistry analysis of NIH/3T3 cells treated with 10µM MG-132 for 6 hours labeling Ubiquitin with Rabbit anti-Ubiquitin antibody (HA750165) at 1/500 dilution.
Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Ubiquitin antibody (HA750165) at 1/500 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.
Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution. -
☑ Cell treatment (CT)
Immunocytochemistry analysis of PC-12 cells treated with 10µM MG-132 for 6 hours labeling Ubiquitin with Rabbit anti-Ubiquitin antibody (HA750165) at 1/200 dilution.
Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Ubiquitin antibody (HA750165) at 1/200 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.
Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution. -
Immunohistochemical analysis of paraffin-embedded human esophagus tissue with Rabbit anti-Ubiquitin antibody (HA750165) at 1/10,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA750165) at 1/10,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX. -
Immunohistochemical analysis of paraffin-embedded mouse esophagus tissue with Rabbit anti-Ubiquitin antibody (HA750165) at 1/10,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA750165) at 1/10,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX. -
Immunohistochemical analysis of paraffin-embedded rat esophagus tissue with Rabbit anti-Ubiquitin antibody (HA750165) at 1/10,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA750165) at 1/10,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX. -
Immunohistochemical analysis of paraffin-embedded mouse brain tissue with Rabbit anti-Ubiquitin antibody (HA750165) at 1/10,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA750165) at 1/10,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX. -
Immunohistochemical analysis of paraffin-embedded rat brain tissue with Rabbit anti-Ubiquitin antibody (HA750165) at 1/10,000 dilution.
The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA750165) at 1/10,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX. -
Flow cytometric analysis of HepG2 cells labeling Ubiquitin.
Cells were fixed and permeabilized. Then stained with the primary antibody (HA750165, 1μg/mL) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"
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Ubiquitin Recombinant Rabbit Monoclonal Antibody [ST46-03]
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