Minimum essential medium-α modification (α-MEM) and fetal bovine serum (FBS) were purchased from Sigma (St. Louis, MO), Penicillin/ Streptomycin from CellGro (Mediatech, Manassas, VA), RANK ligand (RANKL) and MCSF were purchased from R&D Systems (Minneapolis, MN). Leukocyte acid phosphatase kit for tartrate-resistant acid phosphatase (TRAP) staining of osteoclasts, Calcein, LY294002, AMD3100, AKT inhibitor and Mithramycin were purchased from Sigma (St. Louis, MO). Transwell chambers for migration assay were purchased from Corning (Tewksbury, MA). For western blotting analysis, anti-phospho-AKT (Thr308), anti-AKT, anti-phospho GSKα/β Ser21/9, anti-GSK-beta, anti-Actin, anti-Tubulin antibodies were purchased from Cell Signaling Technology (Danvers, MA) and anti-Sp1 antibody, IgG secondary antibody from Millipore (Temecula, CA). HBSS, HEPES and fetal calf serum were purchased from Gibco, Invitrogen (Carlsbad, CA). Rhodamine phalloidin actin stain was purchased from Invitrogen (Carlsbad, CA). Immunofluorescence images were captured using a Leica fluorescent microscope (Buffalo Grove, IL).
Akt inhibitor
Manufactured by Merck Group
Sourced in United States, Germany
The AKT inhibitor is a laboratory reagent used in biochemical and cell biology research. It functions as a selective inhibitor of the AKT protein kinase, which is involved in cellular signaling pathways. The AKT inhibitor can be used to investigate the role of AKT in various cellular processes.
Automatically generated - may contain errors
Lab products found in correlation
Mg132, by Merck Group (2 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
Ly294002, by Merck Group (2 mentions)
Lipofectamine 2000, by Thermo Fisher Scientific (2 mentions)
Turbofect, by Thermo Fisher Scientific (2 mentions)
Ripa lysis buffer, by Thermo Fisher Scientific (2 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (2 mentions)
Mithramycin, by Merck Group (1 mentions)
Fluorescent microscope, by Leica (1 mentions)
Fetal calf serum (fcs), by Thermo Fisher Scientific (1 mentions)
Calcein, by Merck Group (1 mentions)
Hepes, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
Penicillin streptomycin, by Corning (1 mentions)
Rank ligand rankl, by R&D Systems (1 mentions)
Anti tubulin antibody, by Cell Signaling Technology (1 mentions)
Anti actin, by Cell Signaling Technology (1 mentions)
Hanks balanced salt solution (hbss), by Thermo Fisher Scientific (1 mentions)
Amd3100, by Merck Group (1 mentions)
M csf, by R&D Systems (1 mentions)
17 protocols using akt inhibitor
1
Osteoclast Differentiation and Migration
2
Heat Shock and Inhibitor Treatment Protocol
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Cells in pre-warmed FBS-containing medium were exposed to heat shock at 43°C for 2 h in a humidified atmosphere with 5% CO2. Cells were treated with 100 μg/mL cycloheximide (Amresco, Cleveland, OH, USA) or 10 μM MG132 (Sigma Aldrich, St. Louis, MO, USA) for 1 h, or 10 μM Akt Inhibitor (1L6-Hydroxymethyl-chiro-inositol-2-(R)-2-O-methyl-3-O-octadecyl- sn-glycerocarbonate, Sigma Aldrich) for 2 h. Cells were transfected with various plasmid vectors, siRNAs (siGENOME SMARTpool TRIM21 (Horizon Discovery, UK), or Invitrogen Silencer Select siRNAs for AKT1 (S659), CHEK2 (S22120) and RCHY1 (S24704) (Thermo Fisher) and SMARTpool non-targeting siRNA (Horizon Discovery) or Control siRNA (Sigma MissionsiRNA Universal Negative Control) using Lipofectamine2000 or Turbofect (Thermo Fisher) or peiRfect transfection reagent (BioBharati Life Sciences, India). DNA amount for transfection was normalized using empty vector pCDNA3.1.
3
Clonogenic Assay of Single MSCs
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Single MSCs were seeded into individual wells of a 96-well culture plate by limited-dilution assay. Cell suspensions containing 1 × 103 cells in 10 mL complete medium were diluted 1:10 (cells: complete medium), and 100 μL of the dilution (~1 cell/100 μL) was seeded into the 96-well plate. The cells were then cultured under normoxic or hypoxic conditions, or under hypoxia with pretreatment with anti-GRP78 antibody (100 ng/mL) or Akt inhibitor (10−6 M; Sigma-Aldrich, St. Louis, MI, USA) in a humidified incubator. Each well was examined for MSC growth at day 10.
4
Cell Cytotoxicity Assay with AKT Inhibitor
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The cell cytotoxicity assays were conducted using D-Plus CCK-8 (CCK-8-Dongin LS, Seoul, Korea). Before seeding cells, each 96-well plate was coated with 1% gelatin (Sigma, St. Louis, MO, USA) dissolved in 1×phosphate-buffered saline (PBS), incubated for at least 15 min in a CO2 incubator and washed once with 1×PBS. After removing the gelatin, 5,000 cells were seeded per well and incubated overnight. The spent media were replaced with fresh media containing DMSO, DH (Sigma), DH plus 5 μM AKT inhibitor (Sigma) and incubated for 24 h, 48 h. After that, the media were aspirated, and a diluted CCK-8 solution (a 10:1 ratio of media to CCK-8 solution) was added to each well and incubated for 2 h in a CO2 incubator. The absorbance at 450 nm was measured using a microplate reader (Tecan XFluor, Mannedorf, Switzerland).
5
H9c2 Cells Metabolic Modulation
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The H9c2 cell line was purchased from the China Center for Type Culture Collection (CCTCC, China), and the cells were cultured and maintained in Dulbecco’s modified Eagle’s medium (DMEM, HyClone, United States) supplemented with fetal bovine serum (FBS, 10%, Gibco, United States), penicillin (1%, Gibco), and streptomycin at 37°C in a humidified atmosphere (5% CO2 and 95% air). H9c2 cells were incubated with normal glucose (NG, 5.5 mmol/L) or HG (33 mmol/L) for 72 h with or without Met (2.5 mmol/L, Meilunbio, China), a PK2 antagonist (PKRA7, 10 μmol/L, Sigma), and an AKT1/2 kinase inhibitor (AKT inhibitor, 10 μmol/L, Sigma).
6
Glutathione Protects Against Hypoxia-Reoxygenation Injury
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GSH (Sigma-Aldrich, St. Louis, MO, USA) was dissolved in ethanol. An equivalent volume of ethanol (final: 0.01%) or water was added to control and all GSH-containing wells. bEnd.3 cells were exposed to 1 mM GSH for 3 hr before H/R injury (this concentration was chosen to study the cellular protective effect of GSH against H/R injury based on the results of our previous study). The present study consisted of four groups of cells. Normal control (NC) cells were cultured with nontreated media without H/R injury, experimental control (EC) cells were cultured in nontreated medium for 18 hr after 6 hr of H/R injury, and 1 mM GSH cells were pretreated with 1 mM GSH for 3 hr before 6 hr of H/R injury. Cells were then cultured in nontreated medium for 18 hr. Cells in the Akt inhibitor group were treated with 100 nM Akt inhibitor (Sigma-Aldrich, St. Louis, MO, USA) 3 hr before H/R injury.
7
Melatonin Protects bEnd.3 Cells from OGD/R Injury
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Melatonin was purchased from Sigma (Sigma, MO, USA) and dissolved in ethanol. An equivalent volume of ethanol (final: 0.01%) or water was added to control and all Melatonin-containing wells. bEnd.3 cells were exposed to 1–100 nM Melatonin for 24 h before OGD/R injury. The present study consisted of four groups: (1) normal control (NC), bEnd.3 cells cultured with normal media without OGD injury; (2) experimental control (EC), bEnd.3 cells cultured in nontreated medium for 18 h after 6 h of OGD injury; (3) 10 nM Melatonin (Mel 10 nM), bEnd.3 cells treated with 10 nM Melatonin for 24 h before 6 h of OGD injury; these cells were then cultured in nontreated medium for 18 h; (4) 100 nM Melatonin (Mel 100 nM): bEnd.3 cells were also treated with 100 nM Melatonin (100 nM Melatonin group) for 24 h before 6 h of OGD injury. These cells were then cultured in nontreated medium for 18 h. In Akt inhibitor groups, we treated 100 nM Akt inhibitor (Sigma, MO, USA) together with Melatonin.
8
Fucoidan and Akt Inhibitor Modulate HT29 Cell Protein Expression
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Protein expression of HT29 cells following treatment with fucoidan (100 μg/mL) and/or Akt 1/2 kinase inhibitor (10−6 mol/L; Akt inhibitor, Sigma) at indicated time-points was assessed using western blot analysis. Total protein was extracted using RIPA lysis buffer (Thermo Scientific, Rockford, IL, USA). Cell lysates were separated by sodium dodecyl sulfate-polyacryl-amide gel electrophoresis (SDS-PAGE), and proteins were transferred to polyvinylidenefluoride membranes (PVDFs; Millipore, Billerica, MA, USA). The membranes were blocked with 5% skim milk and incubated with primary antibodies against vascular endothelial growth factor (VEGF), phospho-AKT, cyclin-dependent kinase 2 (CDK2), CDK4, Cyclin D, Cyclin E, p21, and β-actin (Santa Cruz Biotechnology, Dallas, TX, USA). After incubation of the membranes with peroxidaseconjugated secondary antibodies (Santa Cruz Biotechnology), bands were visualized using enhanced chemiluminescence reagents (Amersham Biosciences).
9
Gingerol Compounds Modulate Adipogenesis
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Zingerone (88787, >98%), 6-gingerol (G1046, >98%), 8-gingerol (01514, >95%), 10-gingerol (G5798, >95%), d -glucose, 3-isobutyl-1-methylxanthine, dexamethasone, insulin, ethanol (EtOH), Oil Red O, propylene glycol, dimethyl sulfoxide (DMSO), pioglitazone, aldehydes (starting materials for shogaols synthesis), AKT inhibitor, AMPK inhibitor compound C and all other chemicals were of analytical grade and were obtained from Sigma-Aldrich Inc. (Saint Louis, MO, USA). Low glucose Dulbecco’s Modified Eagle Medium (DMEM, low glucose; Catalog No.: 12320), penicillin/streptomycin and fetal bovine serum (FBS), horse serum, trypsin-EDTA were bought from Invitrogen (Carlsbad, CA, USA). Anti-AMPK, anti-phospho-AMPK, anti-aP2, anti-C/EBP-α, anti-FAS, anti-GLUT-4, anti-AKT, anti-phospho-AKT and anti-GAPDH antibodies were purchased from Cell Signaling Technology (Danvers, MA, USA). Anti-SREBP-1 was purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA).
10
Oxidative Stress-Induced Signaling in hMSCs
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Human MSCs (hMSCs) were obtained from American Type Culture Collection (Manassas, VA, USA). Fetal bovine serum (FBS) was purchased from Biowhittaker (Walkersville, MD, USA). Hydrogen peroxide solution was obtained from the Sigma Chemical Company (St. Louis, MO, USA). Phospho-p38 mitogen-activated protein kinase (MAPK), p38 MAPK, phospho-c-Jun N-terminal kinase (JNK), JNK, phospho-ataxia telangiectasia mutated (ATM), ATM, phospho-p53, p53, phospho-PI3K, PI3K, phospho-Akt, and Akt antibodies were obtained from New England BioLabs (Hertfordshire, UK). Manganese superoxide dismutase (MnSOD), Bcl-2, BAX, cleaved caspase-3 (c-caspase-3), and poly (ADP ribose) polymerase-1 (PARP-1) were purchased from Santa Cruz Biotechnology (Delaware, CA, USA). Goat anti-rabbit or mouse IgG antibody was purchased from Jackson ImmunoResearch (West Grove, PA, USA). Lycopene and Akt inhibitor were purchased from Sigma (St. Louis, MO, USA).
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