BJ and HaCaT reporter cells were seeded in white 96-well plates (PerkinElmer) at concentration of 2 × 105 in 100 μL of Minimum Essential Media (Sigma-Aldrich) supplemented by 1% nonessential amino acids (NEAA), 1 mM sodium pyruvate, 10% FCS, streptomycin (100 μg/mL), and penicillin (100 IU/mL) for 24 h at 37°C and 5% CO2. The media was replaced by Opti-MEM media (Gibco) containing 0.5% FBS and 1% NEAA and cells were treated by 40 μg/mL of ginger extract or 30 μM quercetin (Sigma-Aldrich) while the control wells contained media with corresponding concentration of the solvent 0.05% dimethyl sulfoxide (DMSO) as described by Bak et al. [17 (link)]. Following 10 h treatment, the cells were lysed by 20 μL of cell culture lysis buffer (E153A, Promega) and 100 μL of luciferase assay substrate (E1483, Promega) was added. Luminescence was measured by EnSpire multimode plate reader (PerkinElmer). The values from luminescence assays were normalized by values acquired from MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays with the cells that were prepared simultaneously with the same treatment conditions as described by Yamazaki et al. [18 (link)]. All the assays were performed three times with five technical replicates for each treatment.
Minimum essential media
Manufactured by Merck Group
Sourced in United States
Minimum essential media is a cell culture medium designed to provide the basic nutrients required for the growth and maintenance of mammalian cells in vitro. It contains a balanced mixture of amino acids, vitamins, and other essential components to support cell survival and proliferation.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
E1483, by Promega (1 mentions)
White 96 well plate, by PerkinElmer (1 mentions)
Opti mem media, by Thermo Fisher Scientific (1 mentions)
Quercetin, by Merck Group (1 mentions)
Enspire multimode plate reader, by PerkinElmer (1 mentions)
Taurine, by Merck Group (1 mentions)
Non essential amino acids (neaa), by Merck Group (1 mentions)
N1 supplement, by Merck Group (1 mentions)
Dulbecco s modified eagle s media, by Thermo Fisher Scientific (1 mentions)
Transwell filter, by Corning (1 mentions)
Hydrocortisone, by Merck Group (1 mentions)
Neratinib, by Selleck Chemicals (1 mentions)
Afatinib, by Selleck Chemicals (1 mentions)
Snu 216, by Korean Cell Line Bank (1 mentions)
Nci n87, by American Type Culture Collection (1 mentions)
Kato 3, by American Type Culture Collection (1 mentions)
Nugc4, by American Type Culture Collection (1 mentions)
Ocum 1, by Japanese Collection of Research Bioresources Cell Bank (1 mentions)
Snu 16, by American Type Culture Collection (1 mentions)
5 protocols using minimum essential media
1
Ginger Extract and Quercetin Effects on Reporter Cells
2
Imaging of Cultured Kidney and RPE Cells
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Opossum kidney (OK) cells were a gift from Dr. Moshe Levi (UC Denver). Cells were grown on glass coverslips in Dulbecco’s modified Eagle’s media (Life Technologies) supplemented with 10% fetal calf serum (FBS; Life Technologies). Human fetal RPE (hfRPE) cultures were a gift from Dr. Sheldon Miller (National Eye Institute, NIH). The hfRPE cells were grown on Transwell filters (Corning Costar, Corning, NY) coated with human extracellular matrix (BD Biosciences, Franklin Lakes, NJ) in minimum essential media (alpha modification; Sigma Aldrich, St. Louis, MO) supplemented with N1 supplement (Sigma), glutamine-penicillin-streptomycin G (Sigma), non-essential amino acids (Sigma), 250 mg/l taurine (Sigma), 20 µg/l hydrocortisone, (Sigma), 0.013 µg/l triiodothyronine (T3 hormone; Sigma), and 5% FBS [15 (link)]. Cells were fixed with Davidson’s Fixative for 20 min, rinsed three times in PBS, and mounted in PBS for THG/TPAF imaging.
3
Gastric Cancer Cell Line Panel for Targeted Therapies
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Twelve gastric cancer cell lines (ECC10, GCIY, KATO‐III, MKN7, MKN74, NCI‐N87, NUGC3, NUGC4, OCUM‐1, SH‐10‐TC, SNU‐16, and SNU‐216) were used in this study. ECC10, GCIY, and MKN7 were provided by Riken BRC through the National Bio‐Resource Project of Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan. NUGC3, MKN74, and OCUM‐1 were provided by Japanese Collection of Research Bioresources/Health Science Research Resources Bank (JCRB/HSRRB), Osaka, Japan. KATO‐III, NCI‐N87, NUGC4, SNU‐16, SH‐10‐TC were purchased from ATCC (Manassas, VA, USA), and SNU‐216 was obtained from the Korean Cell Line Bank. All the cells, other than GCIY and OCUM‐1, were cultured in RPMI‐1640 media supplemented with 10% FBS. GCIY and OCUM‐1 were cultured in minimum essential media (Sigma‐Aldrich) with 15% FBS and DMEM with 0.5 mmol/L sodium pyruvate and 10% FBS, respectively. Afatinib, neratinib, and PPP were purchased from Selleckchem and MedChem Express. Gefitinib was purchased from Sykkinase. Trastuzumab and pertuzumab were obtained from Chugai Pharmaceutical Co.
4
Xenograft mouse model of breast cancer
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Briefly, L929 cells obtained from National Health Research Institutes were cultured in MEM medium (Minimum Essential Media, Sigma-Aldrich) with 10% Fetal bovine serum (FBS, Gibco) and 1% Antibiotic-Antimycotic (Gibco) and maintained in a 5% CO2 atmosphere at 37 °C.
Breast cancer 4T1 cells obtained from National Health Research Institutes were used as model cancer cells in this study. The 4T1 cells were cultured in RPMI medium (RPMI-1640 Media (Sigma-Aldrich) with 10% Fetal bovine serum (FBS, Gibco) and 1% Antibiotic-Antimycotic (Gibco) and maintained in 5% CO2 atmosphere at 37 °C.
6-week-old male BALB/c nude mice were used as a xenograft animal model in this study. The nude mice were injected subcutaneously with 2 × 106 4T1 cells suspended in 200 μL 1X PBS into the right thigh. All the animal experiments were approved by National Health Research Institutes (NHRI-IACUC-107013).
Breast cancer 4T1 cells obtained from National Health Research Institutes were used as model cancer cells in this study. The 4T1 cells were cultured in RPMI medium (RPMI-1640 Media (Sigma-Aldrich) with 10% Fetal bovine serum (FBS, Gibco) and 1% Antibiotic-Antimycotic (Gibco) and maintained in 5% CO2 atmosphere at 37 °C.
6-week-old male BALB/c nude mice were used as a xenograft animal model in this study. The nude mice were injected subcutaneously with 2 × 106 4T1 cells suspended in 200 μL 1X PBS into the right thigh. All the animal experiments were approved by National Health Research Institutes (NHRI-IACUC-107013).
5
Sampling and Processing of AECOPD Throat Swabs
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Throat swabs were collected from patients with AECOPD within 24 hours of hospital admission, aseptically. The collected swabs were then transported to the laboratory in sterile screw-capped tubes containing minimum essential media (Sigma, USA) supplemented with penicillin (2000 U/mL), gentamicin (1.5 mg/mL), amphotericin B (2 µg/mL) and fetal bovine serum (1.5%). The samples were immediately processed or temporarily stored at 4°C and processed within 48 hours. Blood samples from each patient for the inflammatory marker analysis were collected, serum was separated and stored at -40°C until used.
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