p62/SQSTM1 Antibody (PS00-61)

Novus Biologicals | Catalog # NBP3-32702

Recombinant Monoclonal Antibody
Novus Biologicals
100% Guarantee

Key Product Details

Species Reactivity

Human, Mouse, Rat

Applications

Immunohistochemistry, Immunohistochemistry-Paraffin, Western Blot, Immunocytochemistry/ Immunofluorescence

Label

Unconjugated

Antibody Source

Recombinant Monoclonal Rabbit IgG Clone # PS00-61 expressed in HEK293
View all p62/SQSTM1 Primary Antibodies »

Product Specifications

Immunogen

Synthetic peptide within Human p62/SQSTM1 aa 400 to the C-terminus (Uniprot: Q13501)

Localization

Cytoplasm, Endoplasmic reticulum, Endosome, Lysosome, Nucleus.

Clonality

Monoclonal

Host

Rabbit

Isotype

IgG

Theoretical MW

48 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Scientific Data Images for p62/SQSTM1 Antibody (PS00-61)

p62/SQSTM1 Antibody (PS00-61)

Western Blot: p62/SQSTM1 Antibody (PS00-61) [NBP3-32702]

Western Blot: p62/SQSTM1 Antibody (PS00-61) [NBP3-32702] - Western blot analysis of p62/SQSTM1 on different lysates with Rabbit anti-p62/SQSTM1 antibody (NBP3-32702) at 1/500 dilution.

Lane 1: PC-12 cell lysate (10 ug/Lane)
Lane 2: RAW264.7 cell lysate (10 ug/Lane)
Lane 3: PANC-1 cell lysate (10 ug/Lane)
Lane 4: Mouse liver tissue lysate (20 ug/Lane)
Lane 5: Rat brain tissue lysate (20 ug/Lane)

Predicted band size: 48 kDa
Observed band size: 62 kDa

Exposure time: 2 minutes;

8% 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 (NBP3-32702) at 1/500 dilution was used in 5% NFDM/TBST at room temperature for 2 hours. Goat Anti-Rabbit IgG - HRP Secondary Antibody at 1:300,000 dilution was used for 1 hour at room temperature.
p62/SQSTM1 Antibody (PS00-61)

Immunohistochemistry: p62/SQSTM1 Antibody (PS00-61) [NBP3-32702]

Immunohistochemistry: p62/SQSTM1 Antibody (PS00-61) [NBP3-32702] - Immunohistochemical analysis of paraffin-embedded human colon carcinoma tissue with Rabbit anti-p62/SQSTM1 antibody (NBP3-32702) at 1/400 dilution.

The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) 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 (NBP3-32702) at 1/400 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.
p62/SQSTM1 Antibody (PS00-61)

Immunocytochemistry/ Immunofluorescence: p62/SQSTM1 Antibody (PS00-61) [NBP3-32702]

Immunocytochemistry/ Immunofluorescence: p62/SQSTM1 Antibody (PS00-61) [NBP3-32702] - Immunocytochemistry analysis of A549 cells labeling p62/SQSTM1 with Rabbit anti-p62/SQSTM1 antibody (NBP3-32702) at 1/50 dilution.

Cells were fixed in 4% paraformaldehyde for 10 minutes at 37 ℃, permeabilized with 0.05% Triton X-100 in PBS for 20 minutes, and then blocked with 2% negative goat serum for 30 minutes at room temperature. Cells were then incubated with Rabbit anti-p62/SQSTM1 antibody (NBP3-32702) at 1/50 dilution in 2% negative goat serum overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488) 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.
p62/SQSTM1 Antibody (PS00-61) Western Blot: p62/SQSTM1 Antibody (PS00-61) [NBP3-32702]

Western Blot: p62/SQSTM1 Antibody (PS00-61) [NBP3-32702]

Western blot analysis on different lysates with NBP3-32702 at 1/20,000 dilution. Lane 1: HeLa cell lysate Lane 2: HeLa treated with 50μM Chloroquine for 18 hours cell lysate Lane 3: NIH/3T3 cell lysate Lane 4: C2C12 cell lysate Lane 5: PC-12 cell lysate Lysates/proteins at 20 µg/Lane. Predicted band size: 48 kDa Observed band size: 62 kDa Exposure time: 2 seconds; 4-20% SDS-PAGE gel. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. NBP3-32702 at 1/20,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG [HRP] Secondary Antibody at 1/50,000 dilution was used for 1 hour at room temperature.
p62/SQSTM1 Antibody (PS00-61) Western Blot: p62/SQSTM1 Antibody (PS00-61) [NBP3-32702]

Western Blot: p62/SQSTM1 Antibody (PS00-61) [NBP3-32702]

Western blot analysis on different lysates with NBP3-32702 at 1/5,000 dilution. Lane 1: PANC-1 cell lysate (10 µg/Lane) Lane 2: RAW264.7 cell lysate (10 µg/Lane) Lane 3: Mouse liver tissue lysate (20 µg/Lane) Predicted band size: 62 kDa Observed band size: 62 kDa Exposure time: 50 seconds; 4-20% SDS-PAGE gel. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. NBP3-32702 at 1/5,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG [HRP] Secondary Antibody at 1/50,000 dilution was used for 1 hour at room temperature.

Applications for p62/SQSTM1 Antibody (PS00-61)

Application
Recommended Usage

Immunocytochemistry/ Immunofluorescence

1:50

Immunohistochemistry-Paraffin

1:2000

Western Blot

1:500-1:5000

Formulation, Preparation, and Storage

Purification

Protein A purified

Formulation

PBS (pH7.4), 0.1% BSA and 40% Glycerol

Preservative

0.05% Sodium Azide

Concentration

1 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C short term. Aliquot and store at -20C long term. Avoid freeze-thaw cycles.

Background: p62/SQSTM1

p62/SQSTM1 (ubiquitin-binding protein p62/Sequestrosome-1) is an intracellular protein (theoretical molecular weight 47.7 kDa) which localizes to the cytoplasm, nucleus, autophagosomes and lysosomes in all tissues (1). Structurally, p62/SQSTM1 consists of multiple domains (e.g., Phox1 and Bem1p-PB1, zinc finger-ZZ, TRAF6 binding domain-TB, Keap1-interacting region-KIR, LC3 interacting region-LIR and ubiquitin-associated domain-UBA) for interaction with various protein targets. Additional domains include nuclear export (NES) and nuclear localization signals (NLS1 and NLS2). p62/SQSTM1 is a multifunctional scaffold protein which forms oligomers with itself or with other proteins through interactions with PB1 domains.

Abnormal function of p62/SQSTM1 is associated with a range of disease states such as neurodegeneration, cancer, and metabolic disorders (2). Mutations in the p62/SQSTM1 sequence have been linked to Paget's disease of the bone, amyotrophic lateral sclerosis, and frontotemporal lobar degeneration. In Parkinson's disease, p62/SQSTM1 has been linked to microglia activation and subsequent neuroinflammation (3). Functionally, p62/SQSTM1 is involved in a broad range of cellular processes such as amino acid sensing by interaction with mTORC1, oxidative stress response through interaction with Keap1, and targeting cargo for autophagy by interacting with ubiquitin labeled proteins (1).

To induce selective autophagy, p62/SQSTM1 forms long oligomers or helical filaments which interact with LC3 and ubiquitin labeled proteins and lead to the initiation of the autophagosome formation (2). p62/SQSTM1 is not only a selective autophagy receptor but also an autophagy substrate, as its engulfed by the autophagosome and degraded by the autophagolysosome. Monitoring LC3 levels is the standard for assessing autophagic flux, however monitoring p62/SQSTM1 levels by Western blot in the presence and absence of autophagy inhibitors (e.g., Chloroquine) is also a common practice (4). Besides its activity as a selective autophagy receptor, p62/SQSTM1 also plays a role as an adaptor in signaling cascades leading to NFkB activation downstream of TNF-R, IL-1 beta R, TrkA and p75NTR. Briefly, for NFkB signaling downstream of the TNF-R activation, p62/SQSTM1 engages RIP1 kinase and PKC iota/lambda through the ZZ and PB1 domains, respectively (5).

References

1.Katsuragi, Y., Ichimura, Y., & Komatsu, M. (2015). P62/SQSTM1 functions as a signaling hub and an autophagy adaptor. FEBS Journal. https://doi.org/10.1111/febs.13540

2. Sanchez-Martin, P., & Komatsu, M. (2018). p62/SQSTM1 - Steering the cell through health and disease. Journal of Cell Science. https://doi.org/10.1242/jcs.222836

3. Yao, L., Zhu, Z., Wu, J., Zhang, Y., Zhang, H., Sun, X.,... Lu, G. (2019). MicroRNA-124 regulates the expression of p62/p38 and promotes autophagy in the inflammatory pathogenesis of Parkinson's disease. The FASEB Journal. https://doi.org/10.1096/fj.201900363r

4 Klionsky, D. J., Abdelmohsen, K., Abe, A., Abedin, M. J., Abeliovich, H., Arozena, A. A.,... Zughaier, S. M. (2016). Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition). Autophagy. https://doi.org/10.1080/15548627.2015.1100356

5. Bitto, A., Lerner, C. A., Nacarelli, T., Crowe, E., Torres, C., & Sell, C. (2014). p62/SQSTM1 at the interface of aging, autophagy, and disease. Age. https://doi.org/10.1007/s11357-014-9626-3

Long Name

Sequestosome 1

Alternate Names

A170, EBIAP, ORCA, OSIL, PDB3, Sequestosome 1, SQSTM1, ZIP3

Gene Symbol

SQSTM1

Additional p62/SQSTM1 Products

Product Documents for p62/SQSTM1 Antibody (PS00-61)

Certificate of Analysis

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Product Specific Notices for p62/SQSTM1 Antibody (PS00-61)

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Related Research Areas

Autophagy
NFkB Pathway

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Protocols

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