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AB303543

Anti-Ubiquitin (phospho S65) antibody [EPR24311-222] - BSA and Azide free

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Rabbit Recombinant Monoclonal UBC phospho S65 antibody. Carrier free. Suitable for WB, ICC/IF, Flow Cyt (Intra) and reacts with Human samples.

View Alternative Names

Polyubiquitin-C, UBC

3 Images
Immunocytochemistry/ Immunofluorescence - Anti-Ubiquitin (phospho S65) antibody [EPR24311-222] - BSA and Azide free (AB303543)
  • ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-Ubiquitin (phospho S65) antibody [EPR24311-222] - BSA and Azide free (AB303543)

This data was developed using ab303542, the same antibody clone in a different buffer formulation. Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized PC-3 (human prostate adenocarcinoma epithelial cell) cells labelling Ubiquitin (phospho S65) with ab303542 at 1/100 dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 dilution (Green). Confocal image showing increased cytoplasmic and nuclear staining in PC-3 cells treated with CCCP (30 μM) for 6 h is observed. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue). Secondary antibody only control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 dilution.

Flow Cytometry (Intracellular) - Anti-Ubiquitin (phospho S65) antibody [EPR24311-222] - BSA and Azide free (AB303543)
  • Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-Ubiquitin (phospho S65) antibody [EPR24311-222] - BSA and Azide free (AB303543)

This data was developed using ab303542, the same antibody clone in a different buffer formulation. Flow cytometric analysis of 4% paraformaldehyde fixed 90% methanol permeabilized PC-3 (human prostate adenocarcinoma epithelial cell) treated with 30µM CCCP for 6 hours (Red) / Untreated control (Green) cells labelling Ubiquitin (phospho S65) with ab303542 at 1/50 dilution (1µg) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue). A Goat Anti-Rabbit IgG (Alexa Fluor® 488, ab150081) at 1/2000 dilution was used as the secondary antibody.

Western blot - Anti-Ubiquitin (phospho S65) antibody [EPR24311-222] - BSA and Azide free (AB303543)
  • WB

Supplier Data

Western blot - Anti-Ubiquitin (phospho S65) antibody [EPR24311-222] - BSA and Azide free (AB303543)

This data was developed using ab303542, the same antibody clone in a different buffer formulation. Blocking and diluting buffer and concentration : 5% NFDM/TBST Exposure time : 136 seconds

All lanes:

Western blot - Anti-Ubiquitin (phospho S65) antibody [EPR24311-222] (<a href='/en-us/products/primary-antibodies/ubiquitin-phospho-s65-antibody-epr24311-222-ab303542'>ab303542</a>) at 1/1000 dilution

Lane 1:

Untreated PC-3 (human prostate adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

PC-3 treated with 30 uM CCCP (<a href='/en-us/products/biochemicals/cccp-mitochondrial-oxidative-phosphorylation-uncoupler-ab141229'>ab141229</a>) for 3 hours, whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 8 kDa,200 kDa

false

Exposure time: 136s

Key facts

Host species

Rabbit

Clonality

Monoclonal

Clone number

EPR24311-222

Isotype

IgG

Carrier free

Yes

Reacts with

Human

Applications

ICC/IF, Flow Cyt (Intra), WB

applications

Immunogen

The exact immunogen used to generate this antibody is proprietary information.

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Reactivity data

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Product details

ab303543 is the carrier-free version of ab303542.

Conjugation ready
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. 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.

Compatibility
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.

Patented technology
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

What are the advantages of a recombinant monoclonal antibody?
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 batch production

For more information, read more on recombinant antibodies.

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Properties and storage information

Form
Liquid
Purification technique
Affinity purification Protein A
Storage buffer
pH: 7.2 - 7.4 Constituents: PBS
Shipped at conditions
Blue Ice
Appropriate short-term storage conditions
+4°C
Appropriate long-term storage conditions
+4°C
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Supplementary information

This supplementary information is collated from multiple sources and compiled automatically.

Ubiquitin is a small regulatory protein found in almost all tissues of eukaryotic organisms. It has a molecular weight of approximately 8.6 kilodaltons. It functions mechanically by attaching to proteins through a process called ubiquitination which involves the formation of an isopeptide bond. Ubiquitin molecules can form polyubiquitin chains through different lysine residues such as K48 and K63 that determine their function. These chains label substrate proteins for various fates including degradation. Ubiquitin is expressed ubiquitously in cells reflecting its essential role in maintaining protein homeostasis.
Biological function summary

The ubiquitin system plays a critical role in regulating protein turnover and quality control within cells. It is part of a larger complex known as the ubiquitin-proteasome system (UPS) which is responsible for degrading proteins that need to be turned over. This process is essential for cell cycle control response to oxidative stress and DNA repair. Ubiquitin's role in tagging proteins for degradation or signaling allows cells to respond quickly to changes in their environment and maintain balance.

Pathways

Ubiquitin functions in several important biological pathways including the Wnt and NF-kB pathways. In the Wnt signaling pathway ubiquitination modulates the stability of key components thereby affecting the pathway's overall activity. In the NF-kB signaling pathway ubiquitin labels inhibitor proteins for degradation which releases and activates NF-kB. These pathways highlight ubiquitin's interaction with proteins such as beta-catenin in Wnt and IkB in NF-kB illustrating how it regulates diverse cellular processes.

The dysfunction of the ubiquitin system is linked to neurodegenerative diseases and cancers. Ubiquitin-related defects in protein degradation can lead to the buildup of unwanted proteins contributing to conditions like Parkinson's disease. Connections with cancer are evident as ubiquitin controls cell cycle proteins and aberrant ubiquitination may drive tumor growth and progression. The protein p53 known to be controlled by ubiquitination plays a significant role in cancer related mechanisms when dysregulated. Understanding and targeting ubiquitin-related pathways may provide new therapeutic opportunities for treating these conditions.
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Product protocols

For this product, it's our understanding that no specific protocols are required. You can visit:
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Target data

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 proteotoxic stress response and cell cycle; 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. Linear polymer chains formed via attachment by the initiator Met lead to cell signaling. Ubiquitin is usually conjugated to Lys residues of target proteins, however, in rare cases, conjugation to Cys or Ser residues has been observed. When polyubiquitin is free (unanchored-polyubiquitin), it also has distinct roles, such as in activation of protein kinases, and in signaling. During ubiquitination, the acceptor ubiquitin is positioned in the active site via direct interaction with the E2 ubiquitin-conjugating enzymes such as UBE2R2 (PubMed : 38326650). As a monoubiquitin, its C-terminal glycine is recognized as a C-degron by Cul2-RING (CRL2) E3 ubiquitin-protein ligase complexes (PubMed : 39548056).
See full target information Polyubiquitin-C phospho S65
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Product promise

We are committed to supporting your work with high-quality reagents, and we're here for you every step of the way. In the unlikely event that one of our products does not perform as expected, you're protected by our Product Promise.
For full details, please see our Terms & Conditions

Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.

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