IU1, siRNA and shRNA were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA). MG132 and bortezomib (Velcade) were purchased from Selleckchem (Houston, TX, USA). MTS assay (CellTiter 96 Aqueous One Solution reagent) was purchased from Promega Corporation (Madison, WI, USA). Propidium iodide (PI) and Annexin V-FITC Apoptosis Detection Kit were purchased from Keygen Company (Nanjing, China). Dynabeads antibody coupling kit was from Life technologies. Antibodies used in this study were purchased from following sources: anti-ubiquitin (P4D1) (Santa Cruz Biotechnology); anti-PARP, anti-CDK2, anti-phospho-Rb, anti-Rb, anti-PSA, anti-Bax, anti-GFP, anti-GAPDH (Bioworld Technology, Inc., Louis Park, MN, USA); anti-CDK4, anti-CDK6, anti-phospho-MDM2, anti-P53, anti-USP14, anti-Flag, anti-cyclin D1, anti-p15, anti-p27 (Cell Signaling Technology, Beverly, MA, USA); anti-MDM2 and anti-AR (Abcam, USA).
Anti usp14
Manufactured by Cell Signaling Technology
Sourced in United States
Anti-USP14 is a protein-specific antibody that recognizes the USP14 protein. USP14 is a deubiquitinating enzyme that plays a role in the regulation of proteasomal protein degradation. The antibody can be used to detect and study the USP14 protein.
Automatically generated - may contain errors
Lab products found in correlation
Anti flag, by Cell Signaling Technology (3 mentions)
Anti p27, by Cell Signaling Technology (3 mentions)
Dynabeads antibody coupling kit, by Thermo Fisher Scientific (2 mentions)
Anti cdk4, by Cell Signaling Technology (2 mentions)
Anti cyclin d1, by Cell Signaling Technology (2 mentions)
Anti p53, by Cell Signaling Technology (2 mentions)
Anti ubiquitin, by Cell Signaling Technology (2 mentions)
Anti lc3b, by Cell Signaling Technology (2 mentions)
Anti phospho akt ser473, by Cell Signaling Technology (2 mentions)
Anti gapdh mb001, by Bioworld Technology (2 mentions)
Anti gfp, by Thermo Fisher Scientific (2 mentions)
Anti atp5a, by Abcam (2 mentions)
Anti tom20, by Santa Cruz Biotechnology (2 mentions)
Anti opa1, by Abcam (2 mentions)
Anti phb2, by Santa Cruz Biotechnology (2 mentions)
Anti fis1, by Enzo Life Sciences (2 mentions)
Anti mfn2, by Abcam (2 mentions)
Anti drp1, by Cell Signaling Technology (2 mentions)
Anti hsp60, by Santa Cruz Biotechnology (2 mentions)
Anti mfn1, by Abcam (2 mentions)
8 protocols using anti usp14
1
Comprehensive Protein Analysis Protocol
2
Ubiquitin-mediated protein regulation assay
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Spautin-1 (S7888), Imatinib (S1026), and MG132 (S2619) were from Sellectchem (Houston, TX, USA). SKP2-C25 (M60136) was from Xcessbio Biosciences, Inc. (San Diego, CA). MTS reagent (G3582) was from Promega Corporation (Madison, WI, USA). Co-IP assay kit (14311D) was from Life Technologies (Carlsbad, CA). Antibodies: anti-Ubiquitin (#3936), anti-K63-Ubiquitin (#12930), anti-K48-Ubiquitin (#12805), anti-USP10 (#8501), anti-USP1 (#8033), anti-USP2 (#8036), anti-USP7 (#4833), anti-USP8 (#11832), anti-USP14 (#11931), anti-USP15 (#66310), anti-USP18 (#4813), anti-UCHL1 (#13179), anti-UCHL3 (#8141), anti-CYLD (#8462), anti-A20 (#5630), anti-SKP2 (#2652), anti-p27 (#3686), anti-FLAG (#14793), anti-HA (#3724), anti-phospho-c-Abl(Y245) (#2861), anti-c-Abl (#2862), anti-phospho-STAT5 (#9359), anti-STAT5 (#25656), anti-phospho-Crkl (#3181) and anti-Crkl (#38710) (Cell Signaling Technology, Beverly, MA, USA); anti-UCHL5 (ab124931), anti-USP13 (ab109264) (Abcam, Cambridge, MA); anti-GAPDH (BS60630) (Bioworld Technology, Inc., Louis Park, MN, USA).
3
Antibody Immunoblotting Protocol
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Anti-USP14 (Cell Signaling Technology, #11931), anti-β-actin (MBL, 177-3), anti-HA-tag (MBL, 561), anti-LC3B (Cell Signaling Technology, #2775), anti-p53 (Cell Signaling Technology, #2524), and anti-PrP (Santa Cruz Biotechnology, M20; SPI-Bio, SAF32 and SAF83) antibodies were purchased from the indicated vendors. The anti-PrP mAb132 was a kind gift from Prof. Motohiro Horiuchi (Hokkaido University). Horseradish peroxidase-conjugated anti-goat (Jackson ImmunoResearch), anti-mouse, and anti-rabbit (GE Healthcare Life Sciences) IgG antibodies were used for immunoblotting.
4
Pharmacological Inhibition of Enzalutamide and MK2206 on Prostate Cancer
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IU1 (S7134), enzalutamide (S1250) and MK2206 (S1078) were obtained from Selleckchem (Houston, TX, USA). DMSO was used to dissolve these inhibitors and the inhibitors were stored at − 20 °C. USP14 (sc-76,817) siRNA was obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA). MTS (catalog no. G111) was obtained through Promega Corporation (Madison, WI, USA). Annexin V-FITC/PI apoptosis detection kits of Keygen Company (Nanjing, China)(KGA107) were purchased. Cell Lysis Buffer (#9803) was from Cell Signaling Technology (MA, USA) and stored at − 20 °C. Anti-GAPDH (MB001) was obtained from Bioworld Technology (St.Louis Park, MN, USA). The other antibodies were from Cell Signaling Technology (MA, USA): anti-PARP (#9542), anti-caspase 3 (#9668), anti-cleaved caspase 3 (#9661), anti-caspase 8 (#9746), anti-cleaved caspase 8 (#9496), anti-cleaved caspase 9 (#9501), anti-Bax (#5023), anti-CDK4(#12790), anti-P27(#3686), anti-caspase 9 (#9508), anti-CDK2 (#2546), anti-CyclinD1(#2922), anti-total-AKT (#9272), anti-AR (#5153), anti-USP14 (#11931), anti-Bcl-2 (#15071), anti-GSK3β (#12456), anti-p-GSK3β (Ser9)(#9323), anti-phospho-AKT (Ser473)(#4060), anti-β-catenin (#8480), anti-EGFR (#4267), anti-IGF1R (#9750).
5
Western Blot Antibody Panel for Autophagy
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The commercial antibodies used for Western blot analysis include the following: Anti-phospho-Akt (Ser473) (1:1000 dilution; no. 3787), anti-LC3B (1:1000 dilution; no. 2775), anti-USP14 (rabbit, 1:1000 dilution; no. 11931), anti-Atg14L (1:1000 dilution; no. 5504), and anti-Flag (1:1000 dilution; no. 2368) were from Cell Signaling Technology. Anti-Beclin 1 (1:1000 dilution for Western blot [sc-11427]; 1:200 dilution for immunoprecipitation [sc-48341]), anti-Akt (1:1000 dilution; sc-8312), and anti-USP14 (mouse, 1:100 dilution for immunoprecipitation; sc-393872) were from Santa Cruz Biotechnology. Anti-Myc (1:1000 dilution; 16286-1-AP), anti-HA (1:1000 dilution; 51064-2-AP) anti-Rubion (1:1000 dilution; 21444-1-AP), and anti-GAPDH (1:10,000 dilution; 60004-1-Ig) were from Proteintech. Anti-Tubulin (1:10,000 dilution; PM054), anti-UVRAG (1:1000 dilution; PD027), and anti-p62 (1:5000; PM045) were from MBL. Anti-ubiquitin Lys63-specific antibody (1:2000 dilution; no. 2210353) was from Milllipore. Mouse monoclonal anti-Flag (1:500 for immunoprecipitation; F1804), Anti-Myc, and anti-HA affinity gel (E6779) were from Sigma-Aldrich. IU1 (S7134), MK2206 (S1078), E64D (S7379), rapamycin (S1039), and MG132 (S2619) were from Selleckchem.
6
Mitochondrial Dynamics Protein Analysis
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The following primary antibodies were used: anti‐LC3, anti‐ATP5a, anti‐OPA1, anti‐Mfn1, anti‐Mfn2 and anti‐proteasome 20S (α+β) were from Abcam; anti‐HSP60, anti‐TOM20 and anti‐PHB2 were from Santa Cruz Biotechnology; anti‐DRP1 and anti‐USP14 were from Cell Signaling; anti‐GFP was from Thermo Fisher Scientific; and anti‐Fis1 was from Enzo Life Sciences. HRP‐conjugated anti‐mouse or anti‐rabbit secondary antibodies were from Thermo Fisher Scientific, and Alexa Fluor 488/555‐conjugated anti‐mouse or anti‐rabbit secondary antibodies were from Life Technology. IU1 was purchased from Sigma‐Aldrich. Other sources of the reagents are indicated below when mentioned.
7
Mitochondrial Dynamics Protein Analysis
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In brief, the cells were collected in RIPA buffer (SH‐SY5Y or MEF cells) supplemented with 10 mM NEM, 10 mM MG132 (Sigma) and protease inhibitor cocktail (Roche). The lysate was incubated on ice for 30 min before being centrifuged at 10,000 × g for 15 min at 4°C. 25–40 μg protein was run on SDS gels and transferred to PVDF membranes. The following commercial antibodies were used: anti‐LC3 (1:1,000; Abcam), anti‐PHB2 (1:1,000; Santa Cruz Biotechnology), anti‐HSP60 (1:1,000; Santa Cruz Biotechnology), anti‐ATP5a (1:3,000; Abcam), anti‐actin (1:10,000; Chemicon), anti‐GFP (1:1,000; Invitrogen), anti‐TOM20 (1:5,000; Santa Cruz Biotechnology), anti‐Mfn1 (1:1,000; Abcam), anti‐Mfn2 (1:1,000; Abcam), anti‐USP14 (1:2,000; Cell Signaling), anti‐OPA1 (1:2,000; Abcam), anti‐DRP1 (1:1,000; Cell Signaling) and anti‐Fis1 (1:500; Enzo Life Sciences). For detection, secondary antibodies conjugated to HRP (Chemicon) were used (1:3,000), and immunoreactivity was visualised with ECL chemiluminescence (Amersham).
8
Cholesterol Uptake and Metabolism Assay
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IU1 (S7134), obtained from Selleckchem (Houston, TX, USA), was dissolved into DMSO and stored at −20°C. USP14 siRNA (sc‐76817) was from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Anti‐GAPDH (MB001) was purchased from Bioworld Technology (St. Louis Park, MN, USA). Anti‐CD36 (ab133625), anti‐ABCG1 (ab52617), anti‐SR‐B1 (ab217318) and anti‐SR‐A (ab123946) were from Abcam. Anti‐Ubiquitin and anti‐USP14 were obtained from Cell Signaling Technology (MA, USA). Anti‐ABCA1 was from Santa Cruz, and Anti‐Lox‐1 was from R&D system. Oil Red O was purchased from Sigma‐Aldrich. Human oxidized low‐density lipoprotein (oxLDL) (YB‐002) and human Dil‐labelled oxidized low‐density lipoprotein (Dil‐oxLDL) (YB‐0010) were purchased from Yiyuan Biotechnologies (Guangzhou, China). Dynabeads antibody coupling kit was obtained from Life technologies. PMA (P1585) was from Sigma‐Aldrich. Anti‐CD36 (FITC) (ab39022) was purchased from Abcam.
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