Bovine IFN-γ was purchased from the Kingfisher Group (King-fisher Biotech, Inc., USA). Bafilomycin A1 and E64d were purchased from Abcam (UK). 3-Methyladenine (3-MA) was purchased from Santa Cruz Biotechnology (Santa Cruz Biotechnology Inc., Germany). Chloroquine and rapamycin were purchased from Cell Signaling Technology (USA). The amino acids were purchased from Nanjing Keygen Biotech. Co., Ltd.
Chloroquine
Manufactured by Cell Signaling Technology
Sourced in United States
Chloroquine is a laboratory reagent used for cell signaling research. It functions as an inhibitor of cellular processes, specifically interfering with the activity of enzymes and signaling pathways within cells.
Automatically generated - may contain errors
Lab products found in correlation
Mg132, by Merck Group (2 mentions)
Mg132, by Cell Signaling Technology (2 mentions)
Rapamycin, by Cell Signaling Technology (1 mentions)
Bafilomycin a1, by Abcam (1 mentions)
3 methyladenine 3 ma, by Santa Cruz Biotechnology (1 mentions)
Cycloheximide (chx), by Merck Group (1 mentions)
Lipofectamine 2000 transfection reagent, by Thermo Fisher Scientific (1 mentions)
Antibodies against gapdh, by Cell Signaling Technology (1 mentions)
Antibodies recognizing, by Abcam (1 mentions)
Perifosine, by Selleck Chemicals (1 mentions)
Arsenic trioxide a1010, by Merck Group (1 mentions)
Lysotracker green dnd 26, by Cell Signaling Technology (1 mentions)
Mouse anti β actin antibody, by Proteintech (1 mentions)
Anti phospho mtor ser2448, by Cell Signaling Technology (1 mentions)
Rapamycin, by Abcam (1 mentions)
Lysosome isolation kit, by Merck Group (1 mentions)
Rabbit anti lamp1, by Abcam (1 mentions)
Cycloheximide (chx), by MedChemExpress (1 mentions)
Anti ha, by Cell Signaling Technology (1 mentions)
Anti β actin, by Cell Signaling Technology (1 mentions)
14 protocols using chloroquine
1
Bovine IFN-γ Modulation Assay
2
Cell Culture and Transfection Protocols
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Huh7, SMMC-7721, HCCLM3 and HEK293T cells were obtained from the Cell Bank of the Chinese Academy of Sciences (Shanghai, China) where they were characterized by mycoplasma detection, DNA-Fingerprinting, isozyme detection and cell vitality detection. All cell lines were cultured in DMEM with 10% fetal bovine serum. All these cells were cultured in a humidified incubator at 37 °C in the presence of 5% CO2. Human Cdc5L in pcDNA3.1/myc-His(−) vector was generated according to the standard PCR protocol. All primers and siRNAs sequences were listed in Table S2 . Transfection of plasmids and siRNAs were performed with Lipofectamine® 2000 transfection reagent (Invitrogen, Carlsbad, CA, USA) following the manufacturer’s protocol. Antibodies recognizing Prp19 and Cdc5L were purchased from Abcam Ltd. (Cambridge, UK). Antibodies against GAPDH was from Cell Signaling Technology (Beverly, MA, USA). Chloroquine was dissolved in sterile nuclease-free water (Cell Signaling Technology, Beverly, MA, USA). Cycloheximide was dissolved in sterile nuclease-free water and used at 100 µg/mL (Sigma-Aldrich, St. Louis, MO, USA). MG132 was dissolved in dimethyl sulfoxide (DMSO) and used at 20 µM (Merck KGaA, Darmstadt, Germany).
3
Diverse Autophagy Modulators for Research
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zVAD-fmk (627610) was purchased from Calbiochem (Gibbstown, NJ, USA). Chloroquine (#14774) and LysoTracker Green DND-26 (#8783) were purchased from Cell Signaling (Danvers, MA, USA). 3-Methyladenine (3-MA) (M9281), Bafilomycin-A1 (B1793), Phorbol-12-myristate-13-acetate (PMA) (P8139), Rapamycin (R0395), All-trans retinoic acid (ATRA) (R2625) and arsenic trioxide (A1010) were purchased from Sigma (St Louis, MO, USA). Perifosine (S1037) was purchased from Selleck Chemicals (Houston, TX, USA).
4
Autophagy Regulation via V-ATPase Complexes
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Western blot was utilized to analyze the expression of LC3 and p62/SQSTMI. Rapamycin (100 nm , abcam) and chloroquine (10 µm , Cell Signaling Technology) were used to induce and inhibit autophagy, respectively. The primary antibodies included rabbit anti‐LC3, rabbit anti‐p62/SQSTMI, rabbit anti‐LAMP1 (Abcam) and mouse anti‐β‐actin antibodies (Proteintech), secondary antibodies were goat anti‐rabbit and goat anti‐mouse antibodies (Beyotime). Gray values of protein bands were quantified by ImageJ software.
Related V‐ATPase complexes (V0 and V1 sectors) were also detected by Western blot. IEC‐6 cells were pre‐incubated with bafilomycin A1 (a specific inhibitor of V0‐sector, 100 nm ) for 1 h. The lysosome‐related organelles were separated by a Lysosome Isolation Kit (Sigma‐Aldrich) used for Western blot. The primary antibodies included rabbit anti‐ATP6V0D, anti‐ATP6V1D (Abcam), anti‐Phospho‐mTOR (Ser2448) (Cell Signaling Technology), and mouse anti‐β‐actin antibodies (Proteintech), secondary antibodies were goat anti‐rabbit and goat anti‐mouse antibodies (Beyotime). Gray values of protein bands were quantified by ImageJ software.
Related V‐ATPase complexes (V0 and V1 sectors) were also detected by Western blot. IEC‐6 cells were pre‐incubated with bafilomycin A1 (a specific inhibitor of V0‐sector, 100 n
5
Inhibition of Proteasome and PI3K
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The proteasome inhibitor MG132 (474790) was obtained from Calbiochem EMD Millipore. Chloroquine (14774) was obtained from Cell Signaling Technology. Ammonium chloride (NH4Cl; 254134) was obtained from Sigma-Aldrich. PI3K inhibitors and ubiquitin-activating E1 inhibitor (28 (link)) were provided by the Chemistry Department at Genentech, Inc.
6
Antibody Characterization for Crucian Carp Immunity
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Anti-FLAG (1:5000, catalog no. 14793S), anti-HA (1:5000, catalog no. 3724S), and anti-β-actin (1:5000, catalog number: 4967S) were purchased from Cell Signaling Technology (Danvers, MA) and ABclonal Technology (Wuhan, China), respectively. Anti-TBK1 Ab was generated by immunization of rabbits with a purified peptide corresponding to 360–550 aa of zebrafish TBK1. Anti-CaIRF7 and anti-CaSTAT1 Abs were generated by immunization of rabbits with purified full length peptides of crucian carp IRF7 and STAT1.14 (link),49 (link)
MG132, NH4Cl, Cycloheximide (CHX) and Actinomycin D (ActD) were purchased from MedChemExpress (Monmouth Junction, NJ). Chloroquine was from Cell Signaling Technology (Danvers, MA). Polyinosinic-polycytidylic acid [poly(I:C)] was from Sigma-Aldrich (St. Louis, MO).
MG132, NH4Cl, Cycloheximide (CHX) and Actinomycin D (ActD) were purchased from MedChemExpress (Monmouth Junction, NJ). Chloroquine was from Cell Signaling Technology (Danvers, MA). Polyinosinic-polycytidylic acid [poly(I:C)] was from Sigma-Aldrich (St. Louis, MO).
7
Proteasome and Autophagy Inhibition
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The proteasome inhibitor, MG132(#A2585) was purchased from Stratech Scientific and used at a concentration of 10 μM. The autophagy inhibitor, Chloroquine (#14774) was obtained from Cell Signalling Technology (CST) and used at a concentration of 10 μM. Cycloheximide (#C81040), sourced from Research Products International, was used at 100 μM. The PARP14 catalytic inhibitor (RBN012759) and negative control (RBN250039) were provided by Ribon therapeutics (Cambridge, MA, USA) and used at 0.1 μM (following optimization between 10 nM and 10 μM).26
8
Autophagy Regulation in Cancer Cells
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Anti-ATG5 (C-1; cat. no. sc-133158; 1:500), anti-MAP-LC3-β (G-9; cat. no. sc-376404; 1:500) and anti-sequesteome 1 (SQSTM1; D-3; cat. no. SC-28359; 1:500) primary antibodies were purchased from Santa Cruz Biotechnology, Inc. Anti-β-actin (ACTB; cat. no. 100166-MM10; 1:500) primary antibody was obtained from Sino Biological, Inc., and the anti-clathrin primary antibody was obtained from Covance, Inc. The PCR primers were synthesized by Sangon Biotech Co., Ltd. Atorvastatin calcium was purchased from Sigma-Aldrich (Merck KGaA). Rapamycin and bortezomib were purchased from LC Laboratories. Chloroquine was obtained from Cell Signaling Technology, Inc. EGF was obtained from PeproTech, Inc., and docetaxel from Absin Bioscience Inc. Valproic acid (VPA), brefeldin A, tunicamycin and thapsigargin were obtained from APeXBIO Technology LLC. The Annexin V-FITC apoptosis detection kit (cat. no. AD10) was purchased from Dojindo Molecular Technologies, Inc. All cancer cell lines were purchased from the American Type Culture Collection.
9
Lentiviral Labeling and Transfection of Cancer Cell Lines
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The MDA-MB-231 cells were lentivirally labeled by L2T and MARCH8-GFP (OriGene Technologies, Rockville, MD, USA, Cat RC209891L2) or GFP control using the lentiviruses and labeling protocol previously described [20 (link)]. Transfections of MDA-MB-231, BT-549, and HEK-293 cells were performed using Lipofectamine LTX Reagent (Thermo Fisher Scientific, Waltham, MA, USA, Cat 15338100), Lipofectamine 3000 (Thermo Fisher Scientific, Waltham, MA, USA, Cat L3000001), and FuGENE HD Transfection Reagent (Promega, Madison, WI, USA, Cat E231A), respectively, according to the supplier’s instructions. To block the proteasomal degradation pathway in MDA-MB-231 cells, a 1:1000 dilution of MG-132 (Abcam, Cambridge, UK, Cat ab147047) at a final concentration of 10 µM was used to treat the cells for 6 h to block proteosome degradation pathway. A 1:1000 dilution of chloroquine (Cell Signaling Technology, Danvers, MA, USA, Cat 14774S) at a final concentration of 50 µM was used for 24 h to block the lysosomal degradation pathway.
10
Breast Cancer Cell Lines Characterization
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Human breast cancer cells MDA-MB-231(cat# 300275), MCF-7 (cat# 300273) and T47D (cat# 300353) were purchased from Cell Line Service (CLS)-GmbH. Hs578T (cat# HTB-126) were purchased from ATCC-USA. MDA-MB-231, Hs578T, and MCF-7 were cultured DMEM (Hyclone) and T47D in RPMI (Hyclone). All media were supplemented with 10% fetal bovine serum (Hyclone), 100 U./ml penicillin/streptomycin. All Cells were maintained at 37°C under a humidified atmosphere containing 5% CO2. Carnosol, N-acetylcystein (NAC) were obtained from Sigma Aldrich. 3-methyladenine (3 MA), chloroquine, Bortezomib, and MG-132 were from Cell Signaling. Antibodies to STAT3, phospho-STAT3 (Tyr705), and β-actin were from Santa Cruz Biotechnology.
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