The chemicals used in this study were crizotinib (Selleck Chemicals LLC, Houston, TX, USA), Cyclosporine A (CsA) (J&K chemical Ltd., Beijing, China), PD98059 (Selleck Chemicals LLC, Houston, TX, USA), MK-2206 (Selleck Chemicals LLC, Houston, TX, USA). The primary antibodies against phospho-Erk1/2, Erk1/2, phospho-AKT, AKT, phospho-STAT3, STAT3, phospho-Cdc25c, phospho-CDK1, CDK1, Cyclin B1, Bcl-XL, caspase-3 and PARP were purchased from Cell Signaling Technology (Boston, MA, USA). KSR2 and Cdc25c were obtained from Santa Cruz Biotechnology (Santa Cruz, CA). The secondary antibodies were horse radish peroxidase (HRP)-conjugated anti-rabbit IgG, anti-mouse IgG (Cell Signaling Technology).
Cdc25c
Manufactured by Santa Cruz Biotechnology
Sourced in United States
CDC25C is a protein phosphatase that plays a key role in the regulation of the cell cycle. It is responsible for the dephosphorylation and activation of cyclin-dependent kinases, which are essential for the progression of the cell cycle. CDC25C is a critical component of the cell cycle control machinery and is involved in the transition from the G2 phase to the mitotic phase.
Automatically generated - may contain errors
Lab products found in correlation
Cyclin b1, by Santa Cruz Biotechnology (9 mentions)
Bcl2 associated x (bax), by Santa Cruz Biotechnology (6 mentions)
β actin, by Santa Cruz Biotechnology (5 mentions)
Bcl 2, by Santa Cruz Biotechnology (4 mentions)
Caspase 3, by Santa Cruz Biotechnology (4 mentions)
Bcl xl, by Santa Cruz Biotechnology (4 mentions)
Gapdh, by Santa Cruz Biotechnology (4 mentions)
Cyclin b1, by Cell Signaling Technology (3 mentions)
Polyvinylidene fluoride membrane, by Merck Group (3 mentions)
Cyclin d1, by Santa Cruz Biotechnology (3 mentions)
Cyclin a, by Santa Cruz Biotechnology (3 mentions)
Cleaved caspase 3, by Cell Signaling Technology (3 mentions)
Propidium iodide, by Merck Group (3 mentions)
Bcl xl, by Cell Signaling Technology (2 mentions)
Mus81, by Santa Cruz Biotechnology (2 mentions)
Cyclinb, by Abcam (2 mentions)
β actin, by Abcam (2 mentions)
Rad51, by Abcam (2 mentions)
Bcl 2, by Abcam (2 mentions)
Bcl2 associated x (bax), by Abcam (2 mentions)
27 protocols using cdc25c
1
Signaling Pathways in Cell Cycle Regulation
2
Western Blot Analysis of Cellular Proteins
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Total protein was collected as previously described. Cell lysates were resolved by SDS-PAGE, and proteins were electro transferred to polyvinylidene fluoride membranes (Millipore, USA). The PVDF membranes were blocked with 10% nonfat milk (Solarbio, Beijing, China). The primary antibodies used included MUS81 (1:200 dilution, Santa Cruz, Texas, USA), β-actin (1:3000 dilution, Abcam, MA, USA), pH2AX (1:400 dilution, CST, MA, USA), Cyclin B (1:3000 dilution, Abcam, MA, USA), CHK1 (1:200 dilution, Santa Cruz, Texas, USA), CDC25C (1:200 dilution, Santa Cruz, Texas, USA ) , pCHK1(1:1000 dilution, CST, MA, USA), pCDC25C(1:1000 dilution, CST, MA, USA), RAD51(1:3000 dilution, Abcam, MA, USA), BCL2(1:3000 dilution, Abcam, MA, USA), and BAX (1:3000 dilution, Abcam, MA, USA).
3
DBDFT Anticancer Protocol Evaluation
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DBDFT was synthesized in accordance with the method reported [37] . Fetal bovine serum (FBS), RPMI 1640, penicillin G, streptomycin, and amphotericin B were obtained from Hangzhou Every Green Organism Engineering Materials Co., Ltd (Hangzhou, PR China). Caspase-3, 8 and -9 activity assay and rhodamine 123 detection kits were purchased from NanJing KeyGen Biotech Co., Ltd (Nanjing, PR China). The caspase-9 inhibitor (Z-LEHD-FMK) was purchased from Enzyme Systems (Sacramento, United States). The primers for β-actin, p21, Chk2, Cdc2, Cdc25C, Cyclin B1, p53, Bax, and Bcl-2 were obtained from Shanghai Sangon Biological Engineering Technology and Service Co., Ltd (Shanghai, PR China). Antibodies to β-actin, p21, Chk2, Cdc2, Cdc25C, Cyclin B1, p53, Bax, and Bcl-2 were obtained from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA). The other chemicals used, such as trypsinase, ribonuclease (RNase), methyl thiazolyl tetrazolium (MTT) and propidium iodide (PI), were purchased from Sigma Aldrich Chemical (St. Louis, MO).
4
DNA Damage Response in Rhabdomyosarcoma Cells
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RH30 and RD cells (5 × 104), seeded onto 2% gelatine coated-glass coverslips in 24-well plates, were allowed to attach overnight and then incubated for 48 h in the presence or absence of Olaparib (5 µM) or AZD2461 (10 µM). For IF analysis of the effects induced by the PARPi and IR combination, RH30 and RD cells, pretreated for 24 h with Olaparib or AZD2461 were irradiated and incubated for additional 4 h at 37 °C. IF assays were performed as previously described (Megiorni et al. 2016 (link)) using the following primary antibodies: Cdc2, p-Cdc2, Cdc25C, Cyclin B1, Cyclin D1, RAD51 (1:20 dilution in PBS; Santa Cruz Biotechnology), and γH2AX (1:500 in PBS; Cell Signaling). All single-stained or merged images were acquired with a Zeiss ApoTome microscope (40× magnification) using the Axiovision software (Carl Zeiss, Jena, Germany). For γH2AX and RAD51, focus fluorescence intensity in the respect of cell number in each analysed field was reported.
5
Antioxidant Enzyme Regulation in Cell Culture
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Cell culture reagents were purchased from Sciencell (Carlsbad, CA, USA). The glutathione synthetase (GSS), super oxide dismutase (SOD), glutathione peroxide (GPx), and CDC25C antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). The mouse anti-cyclin B and CDK-1/CDC-2 antibodies were purchased from BD Transduction Laboratories (San Jose, CA, USA). The goat anti-rabbit IgG and goat anti-mouse IgG antibodies were purchased from Santa Cruz Biotechnology. The propidium iodide was purchased from PD bioscience (San Jose, CA, USA). Electrophoresis reagents were purchased from Bio-Rad (Richmond, CA, USA), and nitrocellulose membranes were purchased from Amersham Scientific, Piscataway, NJ, USA. All other reagents were purchased from Sigma–Aldrich (St. Louis, MO, USA).
6
Molecular Mechanisms of ACFP-Mediated Apoptosis
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The ACFP peptide was provided by Shanghai Sangon Biological Engineering Technology, and the purity was confirmed by RP-HPLC to be > 99.5 %. Trizol Kit was purchased from Invitrogen, USA. Reverse Transcription System was purchased from Promega, USA. Eight joint tubes for PCR were purchased from ABI, USA. SYBR Green I Premix Ex Taq was purchased from Takara Biotechnology (Dalian) Co., Ltd, China. Mouse monoclonal antibodies of Bcl-xl, Bax, Akt, Caspase-3, CDC25C, CyclinB1 and β-Actin were purchased from Santa Cruz Biotechnology, Inc. The horseradish peroxidase conjugated secondary antibody (goat anti-mouse IgG) and Super Signal West Pico Trial Kit (ECL chromogenic reagent kit) were purchased from Pierce, USA; BCA Kit for protein quantitative assay was purchased from Shanghai Sangon Biological Engineering Technology.
7
Western Blot Analysis of Cell Signaling Proteins
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EESR-treated cells were lysed with lysis buffer (20 mM Tris-HCl [pH 7.5], 150 mM NaCl, 1 mM ethylenediaminetetraacetic acid, 1 mM ethylene glycol bis(2-aminoethyl ether) tetraacetic acid, 1% Triton X-100, 1 μg/mL leupeptin, 1 mM phenylmethanesulfonyl fluoride) for 1 hour at 4°C and centrifuged for 30 minutes at 13,000 rpm. Total soluble proteins in the supernatant were collected and the concentration of protein was determined by Bradford method. For Western blot analysis, 30 to 50 μg/mL of proteins were resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and blotted onto nitrocellulose membranes. Blots were incubated at 4°C overnight with specific primary antibodies followed by horseradish peroxidase-conjugated secondary antibodies and visualized by an enhanced chemiluminescence detection system (FluoChem®FC2; Alpha-Innotech, San Leandro, CA, USA) using Western blotting luminol reagent (Santa Cruz Biotechnology, Dallas, TX, USA). CDK1, cyclin A, cyclin B, cell division cycle 25C (Cdc25C), p-Cdc25C, Wee1, p53, Fas, Fas-associated protein with death domain (FADD), Bax, caspase-3, caspase-8, caspase-9, PARP, actin primary antibodies and peroxidase-conjugated secondary antibodies were from Santa Cruz Biotechnology. Primary antibodies against CHK2, p-CHK2, p21, p-CDK1, and Bcl-2 were purchased from Cell Signaling Technology (Beverly, CA, USA).
8
Investigating DNA Damage Responses in HepG2 Cells
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Pine needle oil [dissolved in dimethyl sulfoxide (DMSO), ≤0.1%], RNase and propidium iodide (PI) solution were purchased from Sigma-Aldrich (Merck KGaA, Darmstadt, Germany). Anti-γ-H2AX (cat no. 05636; 1:1,000) antibody was purchased from EMD Millipore Billerica, MA, USA), ATM (cat no. 2873; 1:500), p-ATM (cat no. 13050; 1:1,000), p-p53 (S15; cat no. 9286; 1:1,000), p-CDC25C (S216; cat no. 4901; 1:500) p-CHK2 (T68; cat no. 2661; 1:1,000) and CHK2 (cat no. 2662; 1:1,000) antibodies were purchased from Cell Signaling Technology, Inc., (Danvers, MA, USA). CDC25C (cat no. sc-327; 1:1,000), β-actin (cat no. sc-47778; 1:1,000), anti-H2AX (cat no. sc-54606; 1:200), p53 (cat no. sc-98; 1:500) antibodies, goat anti-rabbit (cat no. sc-2030; 1:3,000) and anti-mouse secondary (cat no. sc-2031; 1:3,000) antibodies were purchased from Santa Cruz Biotechnology, Inc., (Dallas, TX, USA). Protein extraction solution kit was purchased from Beijing SBS Genetech Co., Ltd., (Beijing, China). Dulbecco's modified Eagle's medium and bovine serum albumin were purchased from Gibco (Thermo Fisher Scientific, Inc., Waltham, MA, USA). The HepG2 cell line was purchased from the China Center for Type Culture Collection of Wuhan University (Wuhan, China). This cell line was originally thought to be a hepatocellular carcinoma, but is now known to be a hepatoblastoma cell line (21 (link)).
9
Immunoblotting of Whole-Cell Protein Extracts
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Whole-cell protein extracts were prepared with ice-cold RIPA buffer (25 mM Tris-HCl pH 7.6, 150 mM NaCl, 1% NP-40, 1% sodium deoxycholate, 0.1% sodium dodecyl sulfate (SDS)) with Complete Protease Inhibitor Cocktail (Roche Life Sciences, Indianapolis, IN). Each aliquot of protein sample was run on a SDS-polyacrylamide gel electrophoresis and transferred onto a nitrocellulose membrane for immunoblotting with primary antibodies, including ARID1A (Santa Cruz, sc-32761, 1:1000 dilution), CDC25C (Santa Cruz, sc-327, 1:2000 dilution), CDC2 (Santa Cruz, sc-54, 1:2000 dilution), GAPDH (Santa Cruz, sc-365062, 1:5000 dilution), HSP90 (Santa Cruz, sc-69703, 1:5000 dilution), α-tubulin (Santa Cruz, sc-5286, 1:5000 dilution), p-CDC25C (Cell Signaling, #9529s, 1:1000 dilution), p-CDC2 (Cell Signaling, #9111s, 1:1000 dilution), AURKA (Cell Signaling, #14475s, 1:2000 dilution), and cleaved caspase 3 (Cell Signaling, #9661s, 1:2000 dilution) antibodies, followed by horseradish peroxidase-conjugated secondary antibodies (Santa Cruz). Uncropped versions of all blots are shown in Supplementary Figs. 8 -12 .
10
Maduramicin Ammonium Cytotoxicity and Apoptosis
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Maduramicin ammonium (molecular weight = 934.16, purity>97%, by HPLC) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA), dissolved in dimethyl sulfoxide (DMSO) to prepare a stock solution (5 mg/ml), aliquoted and stored at −80°C. Dulbecco's modified Eagle's medium (DMEM) and 0.05% trypsin-EDTA were obtained from Mediatech (Manassas, VA, USA). Fetal bovine serum (FBS) was from Atlanta Biologicals (Lawrenceville, GA, USA). One Solution Cell Proliferation Assay Kit was from Promega (Madison, WI). Cellular DNA Flow Cytometric Analysis Kit was purchased from Roche Diagnostics (Indianapolis, IN, USA). CF488A-Annexin V and Propidium Iodide (PI) Apoptosis Assay Kit was purchased from Biotium (Hayward, CA, USA). Enhanced chemiluminescence solution was from Perkin-Elmer Life Science (Boston, MA, USA). The following antibodies were used: cyclin A, cyclin B1, cyclin D1, cyclin E, CDK1, CDK2, CDK4, CDK6, CDC25A, CDC25B, CDC25C, p21Cip1, p27Kip1, Rb, p-Rb (S807/811), survivin, Mcl-1, Bcl-2, Bcl-xL, BAX, BAK, BAD, FasL, Fas/CD95, TNFα, TNFR1, TRAIL, DR4, DR5, FLIP S/L, FADD, TRADD, RIP (Santa Cruz Biotechnology, Santa Cruz, CA, USA), cleaved caspase 3, cleaved PARP (Cell Signaling, Beverly, MA, USA), β-tubulin (Sigma, St Louis, MO), goat anti-mouse IgG-horseradish peroxidase, and goat anti-rabbit IgG-horseradish peroxidase (Pierce, Rockford, IL, USA).
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