Sqstm1 p62 d5e2

Manufactured by Cell Signaling Technology

Sourced in United States

SQSTM1/p62 (D5E2) is a rabbit monoclonal antibody that recognizes the SQSTM1/p62 protein. SQSTM1/p62 is a multifunctional adapter protein that plays a role in various cellular processes.

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Lab products found in correlation

Benzonase nuclease, by Merck Group (1 mentions) Jbw301, by Merck Group (1 mentions) Brca1 c20, by Santa Cruz Biotechnology (1 mentions) β actin, by Abcam (1 mentions) Eef2k, by Cell Signaling Technology (1 mentions) Hydroxychloroquine sulfate, by Selleck Chemicals (1 mentions) Bafilomycin a1, by Selleck Chemicals (1 mentions) Cleaved caspase 3 asp175, by Cell Signaling Technology (1 mentions) Lc3 1 2, by Cell Signaling Technology (1 mentions) β actin, by Cell Signaling Technology (1 mentions) Mk2206, by Merck Group (1 mentions)

Close spelling variants of this product

P62 (D5E2), SQSTM1/p62

3 protocols using sqstm1 p62 d5e2

Western Blotting Analysis of Autophagy and Nrf2 Signaling

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Human primary synovial fibroblasts and HEK293 cells were collected on ice in lysis buffer [32 (link)]. Human synovial tissue was cut, freezed and macerated in cold lysis buffer. Western blotting was done as explained previously [14 (link), 32 (link)], by the use of following primary antibodies: p62/SQSTM1 (P0067), LC3B (L7543), GAPDH (G9545), α-Tubulin (T9026), Sigma-Aldrich, p62 (PW9860), Enzo Life Sciences, SQSTM1/p62 (D5E2) (#8025), RAD23B (D4W7F) (#13525), Cell Signaling, Phospho-p62 (SQSTM1) (Ser351) (M217-3), MBL, NGLY1 (A305-547A-T) Bethyl Laboratories, Inc., GFP (Y1030), UBPBio, ENGase (PA5-24531), Thermo Fisher Scientific, Nrf2 (MAB3925), R&D, and Nrf2 (Phospho S40) [EP1809Y] (ab76026), Abcam. Rabbit (#7074) and mouse (#7076) secondary antibodies were from Cell Signaling and mouse IgG2b (Star 134) from AbD Serotec.

Relic B., Charlier E., Deroyer C., Malaise O., Crine Y., Neuville S., Gillet P., de Seny D, & Malaise M.G. (2018). Serum starvation raises turnover of phosphorylated p62/SQSTM1 (Serine 349), reveals expression of proteasome and N-glycanase1 interactive protein RAD23B and sensitizes human synovial fibroblasts to BAY 11-7085-induced cell death. Oncotarget, 9(88), 35830-35843.

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Immunoblotting of Nuclear Proteins

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Cell lysates were prepared with 0.5% NP-40 buffer (50 mM Tris–HCl [pH 7.5], 0.5% Nonidet P-40, 150 mM NaCl, 50 mM NaF, 1 mM dithiothreitol, 1 mM NaVO₃, 1 mM PMSF, 2 µg/ml aprotinin, 2 µg/ml leupeptin, 10 µg/ml trypsin inhibitor, and 150 µg/ml benzamidine) and subjected to immunoblotting, as previously described [15 (link), 16 (link)]. Benzonase nuclease (125 U/ml; Millipore, MA, USA) and 2 mM MgCl₂ were supplemented for detection of nuclear proteins. The relative protein expression levels of Trop-2 were measured using a densitometer and normalized to tubulin. The following antibodies were used: rabbit monoclonal antibody against Trop-2 (Abcam, Cambridge, UK), AKT (Cell Signaling Tech, MA, USA), SQSTM1/p62 (D5E2; Cell Signaling Tech), rabbit polyclonal antibody against ERα (Millipore), BRCA1 (C20; Santa Cruz, CA, USA), HER2 (Cell Signaling Tech), pAKT (phospho-Ser473 AKT; Cell Signaling Tech), TFEB (Cell Signaling Tech), pRSK (Pan phospho-Ser221, Ser227, Ser218, Ser232; Invitrogen), LC3 (2775; Cell Signaling Tech), mouse monoclonal antibody against γH2AX (JBW301, Millipore), RSK1 (A-10; Santa Cruz), RSK2 (E-1; Santa Cruz), β-actin (6276; Abcam), and α-tubulin (DM1A; Richard-Allan Scientific, MI, USA).

Zhu J., Wu W., Togashi Y., Taira Nihira N., Johmura Y., Zhu D., Nakanishi M., Miyoshi Y, & Ohta T. (2022). Alteration of Trop-2 expression in breast cancer cells by clinically used therapeutic agents and acquired tamoxifen resistance. Breast Cancer (Tokyo, Japan), 29(6), 1076-1087.

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MK-2206 and Autophagy Inhibitors Assay

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The AKT inhibitor MK-2206 was obtained from Merck & Co., Inc. (Whitehouse Station, NJ, USA). Bafilomycin A1 (Bafil A1), NH125 and hydroxychloroquine sulfate (HCQ) were purchased from Selleck Chemicals (Houston, TX, USA). The working stocks were dissolved to 10 mM, 50 μM and 1 mM using dimethyl sulfoxide (DMSO) for MK-2206, Bafil A1 and NH125, respectively. All reagents were stored at −20°C and diluted in fresh culture medium before experiments. The concentration of DMSO in the final solution did not exceed 1% (v/v). The stock solution of HCQ was formulated to 50 mM with ultrapure water. In vivo, 30% Captisol (CyDex Pharmaceuticals, Inc., Lenexa, KS, USA) was used to dose the MK-2206. LC3 I/II, β-actin, phospho-eEF2 (Thr56), eEF2, eEF2K, SQSTM1/p62 (D5E2) and cleaved caspase-3 (Asp175) were obtained from Cell Signaling Technology (Danvers, MA, USA). Anti-hypoxia-inducible factor-1α antibody was purchased from BD Biosciences (San Diego, CA, USA).

Zhao Y.Y., Tian Y., Liu L., Zhan J.H., Hou X., Chen X., Zhou T., Huang Y, & Zhang L. (2018). Inhibiting eEF-2 kinase-mediated autophagy enhanced the cytocidal effect of AKT inhibitor on human nasopharyngeal carcinoma. Drug Design, Development and Therapy, 12, 2655-2663.

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