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Gp62 c

Manufactured by Novus Biologicals
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The GP62-C is a lab equipment product for general laboratory use. It serves a core function in the scientific research process. No further details are provided to maintain an unbiased and factual approach.

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Lab products found in correlation

Eif2α, by Cell Signaling Technology (1 mentions) P perk, by Bioss Antibodies (1 mentions) α tubulin, by Cell Signaling Technology (1 mentions) P eif2α, by Cell Signaling Technology (1 mentions) P akt, by Cell Signaling Technology (1 mentions) P erk, by Cell Signaling Technology (1 mentions) Oligomycin, by Santa Cruz Biotechnology (1 mentions) Nb100 2220, by Novus Biologicals (1 mentions) Rabbit anti tomm20, by Santa Cruz Biotechnology (1 mentions) Mouse anti atp5b, by Santa Cruz Biotechnology (1 mentions) Antimycin a, by Santa Cruz Biotechnology (1 mentions) Mouse anti parkin, by Cell Signaling Technology (1 mentions) Alexa fluor 647, by Thermo Fisher Scientific (1 mentions) Mitotracker cmxros, by Thermo Fisher Scientific (1 mentions) Alexa fluor 488, by Thermo Fisher Scientific (1 mentions) Bafilomycin a1, by Merck Group (1 mentions) Alexa fluor 594, by Thermo Fisher Scientific (1 mentions) Dimethyl sulfoxide (dmso), by Merck Group (1 mentions) Streptomycin, by Thermo Fisher Scientific (1 mentions) Lysotracker dnd 99, by Thermo Fisher Scientific (1 mentions)

4 protocols using gp62 c

1

Autophagy and Signaling Pathway Analysis

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Freshly harvested or −80°C frozen tissue was ground in liquid nitrogen and lysed using Tris lysis buffer (1 M Tris HCl, 1 M NaCl, 0.1 M EDTA, 10% NP-40). Lysate was run through PAGE (30 μg of protein per well), and blots were probed with antibodies against p62 (Progen, GP62-C), LC3 (Novus Biologicals, NB600-1384), p-PERK (Bioss, BS-3330R), PERK (Cell Signaling, 3192), p-eIF2α (Cell Signaling, 9721), eIF2α (Cell Signaling, 9722), AR, IGF-1β (Cell Signaling, 3027), p-Akt (Cell Signaling, 9271), Akt (Cell Signaling, 9272), p-S6 (Cell Signaling, 4858), S6 (Cell Signaling, 2217), and α-tubulin (Cell Signaling, 2125).
Santanam U., Banach-Petrosky W., Abate-Shen C., Shen M.M., White E, & DiPaola R.S. (2016). Atg7 cooperates with Pten loss to drive prostate cancer tumor growth. Genes & Development, 30(4), 399-407.
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2

Mitochondrial Dysfunction and Autophagy Analysis

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Antimycin A (Santa Cruz), oligomycin (Santa Cruz), CCCP (Sigma), bafilomycin A1 (Sigma), and MitoTracker CMXROS (ThermoFisher) were resuspended in DMSO (Sigma). Antimycin A and oligomycin were stored in aliquots at −80°C. CCCP and bafilomycin A1 were stored in aliquots at −20°C. Primary antibodies for immunofluorescent staining include the following: rabbit anti‐LC3 (Sigma L7543, 1:5,000), rabbit anti‐TOMM20 (Santa Cruz sc‐11415, 1:1,000), mouse anti‐Parkin (Cell Signaling 4211, 1:1,000), mouse anti‐DNA (Millipore CBL186, 1:1,000), mouse anti‐ATP5B (Santa Cruz sc‐166462, 1:1,000), mouse anti‐PEX13 (Santa Cruz sc‐271477, 1:100), rabbit anti‐PMP70 (Thermo Scientific PA1‐650, 1:1,000), mouse anti‐Flag (Sigma 184‐200UG, 1:1,000), and rabbit anti‐WIPI2 (Abcam ab105459, 1:500). Secondary antibodies were conjugated to AlexaFluor488, AlexaFluor594, and/or AlexaFluor647 (Invitrogen, 1:750). Primary antibodies for Western blot analyses include the following: mouse anti‐PEX13 (Santa Cruz sc‐271477, 1:200), rabbit anti‐ATG7 (Sigma A2856, 1:1,000), rabbit anti‐FANCC (Fanconi Anemia Research Fund FANCC‐C2, 1:1,000), mouse anti‐SMURF1 (Sigma WH0057154M1, 1:1,000), guinea pig anti‐p62 (Progen GP62‐C, 1:1,000), rabbit anti‐LC3 (Novus NB100‐2220, 1:1,000), and HRP‐conjugated mouse anti‐actin (Santa Cruz sc‐47778‐HRP, 1:2,000).
Lee M.Y., Sumpter R J.r., Zou Z., Sirasanagandla S., Wei Y., Mishra P., Rosewich H., Crane D.I, & Levine B. (2016). Peroxisomal protein PEX13 functions in selective autophagy. EMBO Reports, 18(1), 48-60.
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3

Osteoclastogenesis Regulation by WDFY3

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All cell incubations were performed in culture medium consisting of αMEM with 2mM L-glutamine, 10% heat-inactivated FBS, 100 IU/ml Penicillin and 100 IU/ml Streptomycin (Life Technologies, 10007D). Mouse soluble RANKL (R&D Systems, 462TR) and M-CSF ELISA (R&D Systems, DY416), were used for in vitro experiments. CMG14-12 (CMG) media was generated as described before [22 ]. Anti-Wdfy3 (Abnova, clone 2F12), anti-WDFY3 (Novus, NBP1-03332), anti-SQSTM1 (Progen, GP62-C), LC3 antibody (Novus, NB100-2220), anti-β-actin antibody (Cell Signaling, 4970), 800 or 680 secondary antibodies (Li-Cor) were used for in vitro experiments. Cyto-ID kit (Enzo biochem, ENZ-51031) was used for autophagosome staining. LysoTracker DND-99 was used for lysosome staining (Life Technologies, L-7528).
Wu D.J, & Adamopoulos I.E. (2017). Loss of WDFY3 Ameliorates Severity of Serum Transfer-Induced Arthritis Independently of Autophagy. Cellular immunology, 316, 61-69.
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4

Protein Extraction and Western Blotting Protocol

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Total cell extracts were prepared in lysis buffer containing 62.5 mM Tris–HCl, 2% (w/v) SDS and 10% sucrose (pH 8) supplemented with EDTA-free protease (Roche, 04693159001) and phosphatase inhibitor cocktails (Roche, 04906837001) and sonicated four times at 60 Hz for 10 s on ice. Fractionation experiments after overexpression of aggregation-prone proteins were performed as described earlier [56 (link)] and specified in the Supplemental Methods. For Western blotting, equal amounts of proteins were separated by SDS-PAGE under denaturing conditions. Proteins were transferred on nitrocellulose membrane and proteins were detected by specific antibodies recognizing SQSTM1 (Progen, GP62-C), LC3B (Novus, NB-100-2220), FLAG (Sigma, F1804), tubulin (Sigma, T9026), SOD1 (Epitomics, 2018-1), ATG9A (Abcam, ab108338), histone H3 (Abcam, ab47915), α-synuclein (Abcam, ab27766), and DNAJC13 (kind gift from M. Farrer, Vancouver, and P. McPherson, Montreal, Canada). Species-specific secondary antibodies coupled to horseradish peroxidase (Dianova) were used to develop the Western blot with the Fusion system (Peqlab) or the Amersham Imager 600 (GE). Densitometry was performed with the AIDA software (Raytest) or ImageJ.
Besemer A.S., Maus J., Ax M.D., Stein A., Vo S., Freese C., Nalbach K., von Hilchen C., Pfalzgraf I.F., Koziollek-Drechsler I., Silva B., Huesmann H., Boukhallouk F., Florin L., Kern A., Behl C, & Clement A.M. (2020). Receptor-mediated endocytosis 8 (RME-8)/DNAJC13 is a novel positive modulator of autophagy and stabilizes cellular protein homeostasis. Cellular and Molecular Life Sciences, 78(2), 645-660.
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