All human oral squamous carcinoma cancer cell lines HSC-3 and OECM-1 were cultured in MEM, RPMI-1640 medium (Gibco, Grand Island, NY, USA), respectively. Medium were supplemented with 10% fetal bovine serum (FBS), 1% non-essential amino acids (NEAA) (Gibco, Grand Island, NY, USA), 0.11 g/L sodium pyruvate (SP) (Applichem, Panreac, Spain), and 1% penicillin/ streptomycin (P/S) (BioInd, Israel) in 5% CO2 at 37 °C. Cell viability was measured by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. HSC-3 and OECM-1 cells (5 × 103 cells per well) were seeded in 96 well-plate and incubated with test compounds for fixed time intervals. Then, the medium was discarded and rinsed by PBS. Afterward, cells were added with 100 μL fresh serum-free containing 10% MTT solution (Cyrusbioscience, Taipei, Taiwan). After incubation at 37 °C for 1 h, MTT-containing solution was removed and the formazan crystals were solubilized with 120 μL of DMSO to detect activities of succinate dehydrogenase in living cells. The absorption levels of each well was measured at 590 nm by a microplate spectrophotometer (Multiskan GO Microplate Spectrophotometer, Thermo Scientific, Vantaa, Finland).
Penicillin streptomycin p s
Manufactured by Sartorius
Sourced in Israel, United States
Penicillin-streptomycin (P/S) is a commonly used antibiotic solution for cell culture applications. It is a combination of the antibiotics penicillin and streptomycin, which work together to inhibit the growth of a wide range of bacteria.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (2 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (2 mentions)
Complete neurobasal medium, by Thermo Fisher Scientific (2 mentions)
Glutamax, by Thermo Fisher Scientific (2 mentions)
Laminin, by Merck Group (2 mentions)
Optiprep, by Merck Group (2 mentions)
Horse serum, by Sartorius (2 mentions)
Brain derived neurotrophic factor (bdnf), by Alomone (2 mentions)
Multiskan go microplate spectrophotometer, by Thermo Fisher Scientific (1 mentions)
Balb 3t3 cells, by Thermo Fisher Scientific (1 mentions)
Poly dl ornithine, by Merck Group (1 mentions)
Insulin, by Merck Group (1 mentions)
Rpmi medium, by Thermo Fisher Scientific (1 mentions)
Puromycin, by Merck Group (1 mentions)
Non essential amino acids (neaa), by Sartorius (1 mentions)
Mda mb 231, by Thermo Fisher Scientific (1 mentions)
Hek293, by Thermo Fisher Scientific (1 mentions)
Dulbecco s modified eagle s medium, by Thermo Fisher Scientific (1 mentions)
Mcf 7, by Thermo Fisher Scientific (1 mentions)
12 protocols using penicillin streptomycin p s
1
Oral Squamous Carcinoma Cell Viability Assay
2
Breast Cancer Cell Culture Protocols
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MCF-7, T47D, BT549 and Hs578T cells were obtained from FuHeng Cell Center, Shanghai, China. All these cells were authenticated by STR profiling by FuHeng Cell Center. LCC2 cells were derived from MCF-7 cells which were continuously exposed to tamoxifen for six months. MCF-7, LCC2, and Hs578T cells were cultured in Dulbecco's modified Eagle's medium (DMEM, BioInd) supplemented with 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin (P/S) (BioInd). T47D and BT549 cells were cultured in RPMI-1640 supplemented with 10% FBS and 1% P/S. All these cells were grown in an incubator at 5% CO2 and humidity.
3
Cell Culture Conditions for HepG2 and BALB/3T3
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HepG2 and BALB/3T3 cells (obtained from the Food Industry Research and Development Institute, Hsinchu, Taiwan) were cultured with DMEM (Invitrogen) supplemented with 10% heat inactivated FBS (Invitrogen) and 1% penicillin/streptomycin (PS; Biological Industries, Beithaemek, Israel) in a humidified atmosphere of 5% CO2 at 37°C. The cells were allowed to grow to ~70% confluence and were split at 1:3 during each passage or used for experiments.
4
Primary Spinal Cord Neuron Isolation
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Primary SC neurons were cultured using E12.5 mouse embryos of either sex as previously described (Zahavi et al., 2015 (link)). Briefly, SCs were excised, trypsinized, and triturated. Supernatant was collected and centrifuged through a 4% BSA cushion. The pellet was resuspended and centrifuged through an OptiPrep gradient (10.4% OptiPrep, Sigma-Aldrich; 10 mm Tricine, 4% glucose) for 20 min at 760 × g with the brake turned off. Cells were collected from the interface, washed once in complete medium, and then plated in coated growth chambers. Cells were maintained in Complete Neurobasal Medium (Invitrogen) containing B27 (Invitrogen), 10% (v/v) horse serum (Biological Industries), 25 nm β-mercaptoethanol, 1% penicillin-streptomycin (PS; Biological Industries), and 1% GlutaMAX (Invitrogen) supplemented with 1 ng/ml GDNF, 0.5 ng/ml CNTF, and 1 ng/ml BDNF (Alomone Labs). Before plating, the growth plates were coated with 1.5 g/ml poly-d l -ornithine (Sigma-Aldrich) overnight at 37°C and 3 g/ml laminin (Sigma-Aldrich) for 2 h at 37°C. For immunofluorescence staining, 30,000 cells were plated on cover slides in 24-well plates. Cells were grown at 37°C in 5% CO2.
5
Cultivation and Glucose Deprivation of Cancer Cell Lines
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The human breast cancer cell lines MCF-7, MDA-MB-231, and Hs-578t, as well as human embryonic kidney cells (HEK293T), were grown in Dulbecco’s Modified Eagle’s Medium (DMEM; Gibco, Grand Island, NY, USA) with 10% fetal bovine serum (FBS; Gibco, Grand Island, NY, USA), 1% penicillin/streptomycin (P/S; Biological Industries, Cromwell, CT, USA), 1% nonessential amino acid (NEAA; Biological Industries, Cromwell, CT, USA), and 1% L-glutamine. An additional 1% insulin (Sigma-Aldrich, St. Louis, MO, USA) was added in the cultured medium for Hs-578t. The MDA-MB-231 cells with a stable knockdown of SIRT3 were grown in DMEM with 1 μg/mL puromycin (Sigma-Aldrich, St. Louis, MO, USA). HCC-1937 was maintained in RPMI medium (Gibco, Grand Island, NY, USA) with 10% FBS and 1% P/S. The cells were cultured in a humidified incubator equilibrated at 37 °C in 5% CO2. For the glucose deprivation experiments, the cells were cultured in glucose-free DMEM (Gibco, Grand Island, NY, USA) or glucose-free RPMI medium (Gibco, Grand Island, NY, USA) with or without glucose (25 mM) (Sigma-Aldrich, St. Louis, MO, USA) for 24 h.
6
Culturing Human Breast and Kidney Cells
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Human breast cancer cell lines were such as MDA-MB-231, MCF-7 and BT474 cells and human embryonic kidney 293 cells (HEK293) were purchased from the Bioresources Collection and Research Center (Hsinchu, Taiwan). All cells were grown in a humidified atmosphere with 5 % CO2 at 37 °C and subcultured with a 0.1 % trypsin, 2 mM EDTA solution. MDA-MB-231, MCF-7 and HEK293 cells were maintained in Dulbecco’s Modified Eagle’s medium (Invitrogen Carlsbad, CA) and BT474 maintained in HybriCare medium (Invitrogen) supplemented with 10 % heat inactivated fetal bovine serum (Gibco BRL, Gaithersburg, USA) and 1 % Penicillin/streptomycin (PS, Biological industries, Beithaemek, Israel).
7
Adipocyte Differentiation Assay Protocol
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Dulbecco’s modified Eagle’s medium (DMEM)/high glucose was obtained from Biological Industries (Israel). Calf serum (CS) for cell culture was purchased from GIBCO BRL (Grand Island, NY, USA). Penicillin/streptomycin (P/S) and fetal bovine serum (FBS) were obtained from Biological Industries (Israel). Rosiglitazone (Rosi), dimethyl sulfoxide (DMSO) and 3-isobutyl-1-methylxanthine (IBMX) were obtained from Sigma-Aldrich (St Louis, MO, USA). Insulin was from Roche (Switzerland), and dexamethasone (DEX) was from Adamas (Switzerland). The 5-amino-4-imidazolecarboxamide ribonucleotide (AICAR) and cell lysis buffer were from Beyotime (Shanghai, China). Compound C was purchased from Calbiochem (San Diego, CA). Bovine serum albumin (BSA) was from Shanghai Sangon Biotech Co., Ltd. (Shanghai, China). The antibodies against AMPK, phospho-AMPK-α (Thr172), FABP4, C/EBPβ, C/EBPα and PPARγ were from Cell Signaling Technology (Beverly, MA, USA), β-actin was from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Triglycerides Kit, Glycerol Assay Kit and Non-esterified fatty acids Kit were purchased from Nanjing Jiancheng Bioengineering Institute. Mouse Adiponectin ELISA Kit was purchased from MULTI SCIENCES.
8
Anticancer Potentials of Purine Derivatives
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Cell culture media, foetal bovine serum, and penicillin-streptomycin (P/S) were purchased from Biological Industries. Vismodegib, cisplatin, etoposide, 5-FU, gemcitabine, 3-(4,5-dymethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), propidium iodide (PI) and dimethyl sulfoxide (DMSO) were obtained from Sigma-Aldrich and AK Scientific. Human colon cancer HCT116 (ATCC® CCL-247EMT™), pancreatic cancer BxPC-3 (ATCC® CRL-1687™) and AsPC-1 (ATCC® CRL-1682™), lung cancer NCI-H1975 (ATCC® CRL-5908™), HEK293 (ATCC® CRL-1573™) and B16F10 (ATCC® CRL-6475™) cells were obtained from ATCC. Medulloblastoma Daoy cells were kindly provided by Dr. Verónica Palma, Universidad de Chile. HT29 cells were obtained from Dr. Juan Villena, Universidad de Valparaíso, Chile. Purine derivatives and control drugs were prepared as fresh DMSO solutions immediately prior to any experiment or stored at -20 °C in amber vials for additional experiments.
9
Isolation and Culture of Primary Spinal Neurons
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Primary spinal cord neurons were cultured using E12.5 mouse embryos of either sex as previously described (Zahavi et al, 2015 (link)). Briefly, spinal cords were excised, trypsinized, and triturated. Supernatant was collected and centrifuged through a 4% BSA cushion. The pellet was resuspended and centrifuged through an Optiprep gradient (10.4% Optiprep (Sigma‐Aldrich), 10 mM Tricine, 4% glucose) for 20 min at 760 g with the brake turned off. Cells were collected from the interface, washed once in complete medium, and then plated in coated growth chambers. Cells were maintained in complete neurobasal medium (Gibco) containing B27 (Gibco), 10% (v/v) horse serum (Biological Industries), 25 nM beta‐mercaptoethanol, 1% penicillin–streptomycin (PS; Biological Industries), and 1% GlutaMAX (Gibco) supplemented with 1 ng/ml glial‐derived neurotrophic factor (GDNF), 0.5 ng/ml ciliary neurotrophic factor (CNTF), and 1 ng/ml brain‐derived neurotrophic factor (BDNF) (Alomone Labs). Prior to plating, growth plates were coated with 1.5 g/ml poly d ‐l ‐ornithine (PLO; Sigma‐Aldrich) overnight at 37°C and with 3 µg/ml Laminin (Sigma‐Aldrich) for 2 h at 37°C. For immunofluorescence staining, 10,000 cells were plated on cover slides in 24‐well plates. Cells were grown at 37°C in 5% CO2.
10
Anti-inflammatory Potential of TCM Extracts
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Human acute monocytic leukemia (THP-1) cells (BCRC, Hsinchu, Taiwan) were cultured in RPMI-1640 medium (Gibco BRL, Gaithersburg, MD, USA) supplemented with 0.05 mM β-mercaptoethanol (Sigma-Aldrich, St. Louis, MO, USA), 2 g/L sodium bicarbonate, 1% penicillin/streptomycin (PS, Biological Industries, Beithemeek, Israel), and 10% fetal bovine serum (Gibco BRL, Gaithersburg, MD, USA) in a humidified atmosphere with 5% CO2 at 37°C. For anti-inflammatory test, THP-1 cells were seeded into 24-well culture plates at 1 × 105 cells/mL/well, and then 1 μg/mL lipopolysaccharides (LPS) (Sigma-Aldrich, St. Louis, MO, USA) were added to the wells to induce inflammation. The TCM extracts, AE, RE, or ARE (final concentration = 0.2%), were then added to the wells. After 24 h of coincubation, the supernatants were obtained by centrifuged at 15,000 ×g for 15 min and assayed using human TNF-α enzyme-linked immunosorbent assay (ELISA) kits (R&D System, MN, USA).
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