MCF-7, a human breast adenocarcinoma cell line, was cultured in an RPMI 1640 medium (Nacalai Tesque; #30264-85) supplemented with 10% fetal calf serum (FCS) and 1% penicillin-streptomycin mixed solution (Nacalai Tesque; #09367-34). MCF10A, a human non-tumorigenic epithelial cell line, was cultured in a DMEM/Ham’s F-12 medium with l -Glutamine, sodium pyruvate, and HEPES without phenol red (Nacalai Tesque; #05177-15) supplemented with 5% FCS, 10 μg/mL insulin, 0.5 μg/mL hydrocortisone, 0.1 μg/mL cholera toxin (Sigma Aldrich, St. Louis, MO, USA; #C8052), 20 ng/mL epidermal growth factor, and 1% penicillin-streptomycin. These cells were cultured at 37 °C under humidified 5% CO2 conditions.
Dmem ham s f 12 medium
Manufactured by Nacalai Tesque
Sourced in Japan
DMEM/Ham's F-12 medium is a cell culture medium commonly used for the in vitro cultivation of a variety of cell types, including mammalian cells. It is a combination of Dulbecco's Modified Eagle's Medium (DMEM) and Ham's F-12 nutrient mixture, providing a balanced formulation of amino acids, vitamins, salts, and other components necessary for cell growth and maintenance.
Automatically generated - may contain errors
Lab products found in correlation
Penicillin streptomycin, by Nacalai Tesque (4 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions)
Dmem with high glucose, by Nacalai Tesque (2 mentions)
Penicillin streptomycin, by Merck Group (1 mentions)
Hydrocortisone, by Merck Group (1 mentions)
Cholera toxin, by Merck Group (1 mentions)
Rpmi 1640 medium, by Nacalai Tesque (1 mentions)
Enzymatic dissociation kit, by STEMCELL (1 mentions)
Basic fibroblast growth factor (bfgf), by Thermo Fisher Scientific (1 mentions)
Epidermal growth factor (egf), by Thermo Fisher Scientific (1 mentions)
Transferrin, by Fujifilm (1 mentions)
Sodium selenite, by Fujifilm (1 mentions)
Insulin, by Fujifilm (1 mentions)
Kanamycin, by Fujifilm (1 mentions)
Tnf α, by R&D Systems (1 mentions)
Penicillin, by Nacalai Tesque (1 mentions)
Streptomycin, by Nacalai Tesque (1 mentions)
Bronchialife epithelial basal medium, by Lifeline Cell Technology (1 mentions)
Lifefactors kit, by Lifeline Cell Technology (1 mentions)
Rpmi 1640, by Nacalai Tesque (1 mentions)
9 protocols using dmem ham s f 12 medium
1
Culturing MCF-7 and MCF10A Cell Lines
2
Isolation of Mouse Neural Stem Cells
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Primary neural stem/progenitor cells (NSPCs) were isolated from the cerebra of B6C3F1 mice and C57BL/6N mice (Japan SLC; Hamamatsu, Japan) using an enzymatic dissociation kit (StemCell Technologies, Vancouver, BC, Canada). The care and use of animals in this study complied with the relevant laws and institutional guidelines for animal welfare at Osaka Prefecture University. For isolation of mouse NSPCs, the striata or the brain tissues of the subventricular zone were minced using a pair of scissors until no large pieces remained, and incubated in the dissociation solution for 7 min at 37°C, followed by treatment with the inhibition solution. The cell suspension was centrifuged at 1200 rpm for 5 min at room temperature, and the cell pellet was washed three times with the resuspension solution. Then, the cells were cultured in DMEM/Ham’s F-12 medium (Nacalai Tesque, Kyoto, Japan) supplemented with 2% B-27 (Thermo Fisher Scientific, Waltham, MA), 20 ng/ml basic fibroblast growth factor (bFGF) (PeproTech, Rocky Hill, CT), 20 ng/ml epidermal growth factor (EGF) (PeproTech), 2 μg/ml heparin sodium salt, and 1% penicillin–streptomycin (Nacalai Tesque) as floating neurospheres at 37°C under humidified 5% CO2 conditions.
3
ATDC5 Chondrocyte Differentiation Assay
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ATDC5 mouse chondrogenic cell line was maintained in DMEM/Ham’s F12 medium (Nacalai Tesque, Kyoto, Japan) supplemented with 5% fetal bovine serum (Thermo Fisher Scientific, Waltham, MA, USA), 100 U/ml kanamycin (FUJIFILM Wako Pure Chemical, Osaka, Japan) in a humidity incubator at 37 °C with 5% CO2. For the induction of chondrocyte differentiation, insulin (10 μg/ml), transferrin (10 μg/ml), and sodium selenite (3 × 10–8 M) (FUJIFILM Wako Pure Chemical) were added to the medium8 (link),9 (link).
Cells were plated on a 12-well plate at a density of 4 × 104 cells/well and used 14–19 days after the induction of chondrocyte differentiation. The medium was changed every other day. Differentiated ATDC5 cells were cultured with KUS121 (0 [DMSO], 50, 100 μM) in the presence of TNF-α (R&D Systems, Minneapolis, MN, USA, 10, 20 ng/ml) or TM (Nacalai Tesque, 0.2 or 3 μg/ml) or under no glucose conditions.
Cells were plated on a 12-well plate at a density of 4 × 104 cells/well and used 14–19 days after the induction of chondrocyte differentiation. The medium was changed every other day. Differentiated ATDC5 cells were cultured with KUS121 (0 [DMSO], 50, 100 μM) in the presence of TNF-α (R&D Systems, Minneapolis, MN, USA, 10, 20 ng/ml) or TM (Nacalai Tesque, 0.2 or 3 μg/ml) or under no glucose conditions.
4
Cell Culture Protocols for Cancer Research
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HEK293T cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM; Nacalai Tesque, Cat# 08459-64). RPE-1 cells (a kind gift from Dr. Hochegger) and derived cell lines were cultured in DMEM-Ham’s F-12 medium (Nacalai Tesque, Cat# 11581-15). H3122 (a kind gift from Dr. Kobayashi), H1975, H1819, H358, A427 and H2009 cells were cultured in RPMI 1640 (Nacalai Tesque, Cat# 30264-56). These media were supplemented with 10% fetal bovine serum (Gibco), 100 U/ml penicillin and 100 μg/ml streptomycin (Nacalai Tesque, Cat# 26252-94). SAEC (human small airway epithelial cells, a kind gift from Dr. Kiyono) were immortalized via the expression of hTERT, a CDK4 mutant and cyclin D1. SAECs and derived cell lines were cultured in BronchiaLife™ Epithelial Basal Medium (Lifeline Cell Technology, Cat# LM-0007) supplemented with the components of a LifeFactors® Kit (Lifeline Cell Technology, Cat# LS-1047, containing human serum albumin, lecithin, linoleic acid, L-glutamine, bovine pituitary extract, and TM-1 factor). All cells were cultured at 37 °C in 5% CO2.
5
Establishment of IFT27 and CEP19 Knockout Cell Lines
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Antibodies, chemicals, and plasmids used in this study are listed in Supplemental Table S1. HEK293T cells (RBC2202; RIKEN BioResource Research Center) were cultured in DMEM with high glucose (Nacalai Tesque) supplemented with 5% fetal bovine serum (FBS). hTERT-RPE1 cells (CRL-4000; American Type Culture Collection) were grown in DMEM/Ham’s F-12 medium (Nacalai Tesque) supplemented with 10% FBS and 0.348% sodium bicarbonate at 37°C in 5% CO2. IFT27-KO cells (cell line #IFT27-2-2) and CEP19-KO cells (cell lines #CEP19-1-2 and #CEP19-1-12) were established from hTERT-RPE1 cells, as described previously (Nishijima et al., 2017 (link); Zhou et al., 2022 (link)).
6
Culturing Human Oral Squamous Cell Carcinoma
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The human oral squamous cell carcinoma cell lines used in this study, H103, H400, H413, H357, H376 and H314, were kindly provided by Professor. Dr. Ian Charles Paterson (University of Malaya). In addition, 3T3 (normal mouse fibroblast) cells were used as normal cells. OSCC cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM)/Ham’s F-12 medium (Nacalai Tesque, Kyoto, Japan) supplemented with 10% fetal bovine serum (J R Scientific, Inc., USA), 100 Units/mL penicillin and 100 μg/mL streptomycin (Sigma-Aldrich) at 37ºC in a humidified atmosphere of 5% CO2. Growth and morphology of the cells were regularly monitored and the medium renewal was done every 2 to 3 days until the cells were confluent.
7
Characterization of human IFT54/TRAF3IP1 isoforms
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IFT54 constructs used in this study are listed in Table S1 . In the current and previous studies (Hiyamizu et al., 2023 (link); Katoh et al., 2016 (link)), we used a cDNA encoding human IFT54/TRAF3IP1 isoform 2 (625-amino acids; NP_001132962) instead of isoform 1 (691-amino acids; NP_056465), because the IFT54 proteins from most other vertebrate species correspond to isoform 2. The other constructs were described previously (Hiyamizu et al., 2023 (link)). Antibodies used in this study are listed in Table S2 . Glutathione S-transferase (GST)-tagged anti-mCherry Nb (LaM-2 version) prebound to glutathione–Sepharose 4B beads were prepared as described previously (Ishida et al., 2021 (link); Katoh et al., 2015 (link)). SAG was purchased from Enzo Life Sciences. hTERT-RPE1 cells (American Type Culture Collection, CRL-4000) were grown in DMEM/Ham's F-12 medium (Nacalai Tesque) supplemented with 10% fetal bovine serum (FBS) and 0.348% sodium bicarbonate at 37°C in 5% CO2. HEK293T cells (RIKEN BioResource Research Center, RBC2202) were cultured in DMEM with high glucose (Nacalai Tesque) supplemented with 5% FBS.
8
Immortalized Mesangial Cell Culture
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A mouse immortalized mesangial cell line, SV40MES13 (MES13), was purchased from the American Type Culture Collection (Manassas, VA, USA). Cells were cultured in a 3:1 mixture of Dulbecco’s modified Eagle’s medium(DMEM)/Ham’s F12 medium (Nacalai Tesque, Kyoto, Japan) with supplemented 5% fetal bovine serum (FBS) (Corning Life Sciences, NY, USA), 14 mM HEPES (Gibco, MD, USA), and 100 U/mL penicillin/streptomycin (Nacalai Tesque). Cells were incubated in a humidified incubator at 37 °C with 5% CO2. All experiments were performed between passages 8 and 9 to minimize the effects of phenotypic variation in continuous culture. Cells were serum-starved with 1% FBS for 24 h or 48 h prior to examination.
9
Harmine and Smoothened Agonist Modulate Lung Explants
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The E11.5 lungs were dissected from WT and Dyrk2−/− mice. The lungs were placed on a Transwell polyester membrane cell culture insert (Corning) and cultured at the air liquid interface in DMEM/ Ham’s F12 medium (Nacalai tesque) supplemented with 10% FBS, penicillin-streptomycin (Nacalai tesque), and Amphotericin B (Sigma) with or without 50 μM Harmine (Tokyo Chemical Industry Co., Ltd.) or 14.8 nM smoothened agonist (SAG) (Enzo Life Sciences). DMSO was used as a diluent control. After 24 or 48 h incubation, the lung explants were collected and used for further analysis.
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