Fetal bovine serum (fbs)

Manufactured by Funakoshi

Sourced in Japan

Fetal Bovine Serum (FBS) is a common laboratory reagent derived from the blood of bovine fetuses. It provides a complex mixture of proteins, growth factors, and other nutrients that support the growth and maintenance of various cell types in cell culture systems. FBS serves as a supplement to cell culture media, promoting cell proliferation and survival.

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10 protocols using fetal bovine serum (fbs)

Cell Adaptability to Metallic Vessels

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MCF-7 (RIKEN BRC, Saitama, Japan) and NHDF cells (Cosmo Bio Co., Ltd., Tokyo, Japan) were cultured in Dulbecco’s modified Eagle’s medium (11965092; Thermo Fisher Scientific Inc.) supplemented with 10% fetal bovine serum (Funakoshi Co., Ltd., Tokyo, Japan) and 1% penicillin (15140122; Thermo Fisher Scientific Inc.) at 37 °C under 5% CO₂. Cells were detached using 0.05% trypsin-ethylenediaminetetraacetic acid (25300; Life Technologies, Carlsbad, CA, USA). Except for the experiments, the cells were cultured in conventional plastic culture dishes.
To demonstrate the cell adaptability (Fig. 2E to H), 1.0 × 10⁶ cells were seeded into the metallic culture vessels and cultured for 24 h, while cells with the same density on culture surfaces were seeded and cultured in 35-mm plastic culture dishes. After culturing, the supernatant was collected to count the number of cells detached from the culture surface. To measure cell viability, live and dead cells were stained with calcein-AM and propidium iodide, respectively. The ratios of live-cell areas were used to determine cell viability.

Imashiro C., Jin Y., Hayama M., Yamada T.G., Funahashi A., Sakaguchi K., Umezu S, & Komotori J. (2023). Titanium Culture Vessel Presenting Temperature Gradation for the Thermotolerance Estimation of Cells. Cyborg and Bionic Systems, 4, 0049.

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Maintaining Alphoid^tetO-HAC Cell Lines

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Chicken B-lymphoma DT40 cells
containing the alphoid^tetO-HAC were cultured in Roswell
Park Memorial Institute medium 1640 (RPMI-1640, GIBCO) supplemented
with 10% fetal bovine serum (Biowest, Nuaille, France), 1% chicken
serum (Invitrogen), 50 μM 2-mercaptoethanol (SIGMA) and 15 μg/mL
Blasticidin S (Funakoshi, Tokyo, Japan) at 40 °C in 10% CO₂. Hypoxanthine phosphoribosyltransferase (HPRT)-deficient
Chinese hamster ovary (CHO) cells (JCRB0218) carrying the alphoid^tetO-HAC were maintained in Ham’s F-12 nutrient mixture
(Invitrogen, USA) plus 10% fetal bovine serum (FBS) with 8 μg/mL
Blasticidin S Hydrochloride (Funakoshi, Japan). Blasticidin S is required
for stable propagation of alphoid^tetO-HAC in the medium.

Lee N.C., Kim J.H., Petrov N.S., Lee H.S., Masumoto H., Earnshaw W.C., Larionov V, & Kouprina N. (2017). Method to Assemble Genomic DNA Fragments or Genes on Human Artificial Chromosome with Regulated Kinetochore Using a Multi-Integrase System. ACS Synthetic Biology, 7(1), 63-74.

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Culturing Drosophila OSCs in Supplemented Medium

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OSCs were obtained from fGS/OSS^{49 (link)}, and cultured at 27 °C in Shields & Sang M3 Insect Medium (Sigma-Aldrich), supplemented with 10% fly extract^{49 (link)}, 10% fetal bovine serum (Funakoshi), 1% l-glutathione reduced (Sigma-Aldrich), and 1% human recombinant insulin (Wako).

Yamaguchi S., Oe A., Nishida K.M., Yamashita K., Kajiya A., Hirano S., Matsumoto N., Dohmae N., Ishitani R., Saito K., Siomi H., Nishimasu H., Siomi M.C, & Nureki O. (2020). Crystal structure of Drosophila Piwi. Nature Communications, 11, 858.

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Culturing Murine Colon Cancer CT26 Cells

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Murine colon cancer CT26 cells (American Type Culture Collection) were maintained in 100 mm tissue culture dishes and cultured in RPMI1640 (FUJIFILM Wako Pure Chemical) medium containing 10% fetal bovine serum (FBS; Funakoshi) and 1% penicillin–streptomycin solution (Fujifilm Wako Pure Chemical) under 5% CO₂ at 37°C.

Kaneko M., Yamazaki H., Ono T., Horie M, & Ito A. (2023). Effective magnetic hyperthermia induced by mitochondria‐targeted nanoparticles modified with triphenylphosphonium‐containing phospholipid polymers. Cancer Science, 114(9), 3750-3758.

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Cell culture protocol for MCF-7 and NHDF

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MCF-7 cells (RIKEN BRC, Saitama, Japan) and NHDFs (Cosmo Bio Co., Ltd., Tokyo, Japan) were cultured in Dulbecco’s Modified Eagle’s Medium (DMEM) (11965092; Thermo Fisher Scientific Inc.) supplemented with 10% fetal bovine serum (FBS) (Funakoshi Co., Ltd., Tokyo, Japan) and 1% penicillin (15140122; Thermo Fisher Scientific Inc.) at 37 °C under 5% CO₂. Cell detachment was performed using 0.05% Trypsin–EDTA (25300; Life Technologies, Carlsbad, CA, USA).

Imashiro C., Takeshita H., Morikura T., Miyata S., Takemura K, & Komotori J. (2021). Development of accurate temperature regulation culture system with metallic culture vessel demonstrates different thermal cytotoxicity in cancer and normal cells. Scientific Reports, 11, 21466.

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CT26 Cell Culture Protocol

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CT26 cells were purchased from the American Type Culture Collection (ATCC). CT26 cells were seeded into 100‐mm dishes with 1.0 × 10⁶ cells/dish in 10 ml of RPMI 1640 (FUJIFILM Wako Pure Chemical) supplemented with 1% penicillin–streptomycin (FUJIFILM Wako Pure Chemical) and 10% FBS (Funakoshi), and cultured at 37°C in 5% CO₂.

Kaneko M., Yamaguchi A, & Ito A. (2022). Induction of immunogenic cell death in murine colon cancer cells by ferrocene‐containing redox phospholipid polymers. Cancer Science, 113(10), 3558-3565.

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Mouse Mammary Carcinoma Metastasis Model

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C3H/He mouse mammary carcinoma (FM3A‐Luc) cells, which express the luciferase gene²⁷ were used. Cells were cultured in 10% (v/v) FBS (Funakoshi), to which 1% (v/v) L‐glutamine‐penicillin–streptomycin (Sigma‐Aldrich) and 0.5 mg/ml G418 (Sigma‐Aldrich) was added. The medium was changed every 4–5 days. Cells were used in experiments after three passages. Lack of Mycoplasma contamination was confirmed on the inoculation day (MycoAlert Mycoplasma Detection Kit; Lonza Rockland). FM3A‐Luc cells (3.3 × 10⁵ cells/ml; 19,800 cells/60 μl), suspended in a mixture of 20 μl PBS and 40 μl of 400 mg/m Matrigel (Collaborative Biomedical Products), were injected into the SiLN to induce metastasis in the PALN and lungs.

Sora S., Sukhbaatar A., Fukushige S., Mori S., Sakamoto M, & Kodama T. (2022). Combination therapy of lymphatic drug delivery and total body irradiation in a metastatic lymph node and lung mouse model. Cancer Science, 114(1), 227-235.

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Characterization of MT-9 Cell Line

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A cultured cloned cell line, MT-9, was used; MT-9 was derived from a spontaneously occurring rat MFH [3 (link)]. MT-9 cells have been subcultured in Dulbecco’s modified Eagle’s medium (DMEM) (Thermo Fisher Scientific Inc., Waltham, MA, USA) added with 10% fetal bovine serum (FBS) (Funakoshi Co. Ltd., Tokyo, Japan) in a Nunc EasYFlask 25 cm² or 75 cm² (Thermo Fisher Scientific Inc., Waltham, MA, USA) at 37 °C and 5% CO₂ atmosphere. MT-9 cells were used for analyses of A3-recognizing epitopes.

Katou-Ichikawa C., Nishina H., Tanaka M., Takenaka S., Izawa T., Kuwamura M, & Yamate J. (2020). Participation of Somatic Stem Cells, Labeled by a Unique Antibody (A3) Recognizing Both N-glycan and Peptide, to Hair Follicle Cycle and Cutaneous Wound Healing in Rats. International Journal of Molecular Sciences, 21(11), 3806.

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Regulation of GLP-1 Secretion in STC-1 Cells

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STC-1 cells (mouse enteroendocrine cell line; ATCC) were cultured in Dulbecco's modified eagle medium (DMEM; Sigma, St. Louis, MO, USA) containing 1% penicillin and streptomycin (Invitrogen), 5% fetal bovine serum (FBS; Funakoshi, Tokyo, Japan), and 15% horse serum (Gibco, NY, USA). To measure GLP-1 secretion, the cells were plated in 24-well-plates (approximately 1 × 10⁵ cells/well) and cultured for 48 h (17 (link)). The cells were treated with octanoate (C8:0), decanoate (C10:0), dodecanoate (C12:0), and embelin (Cayman Chemical, MI, USA) for 1 h. The culture supernatant was collected in the presence of a DPP-IV inhibitor. For siRNA-mediated knockdown, STC-1 cells were transfected with 30 nM siRNA (universal control: 5′-UGGUUUACAUGUCGACUAA-3′, Gpr84: 5′-GUUGGGCUAUCGAUACUUU-3′, Dharmacon, Lafayette, CO, USA) using Lipofectamine 2000 transfection reagent (Invitrogen), following the manufacturer's instructions. For inhibitor treatment, STC-1 cells were pretreated with the Gα(i/o) blocker NF023 (10 μM; Calbiochem, CA, USA), Gi/o-type G protein inactivator pertussis toxin (PTX; 1 μg/mL; Wako), Gβγ blocker Gallein (30 μM; Merck Millipore), the phosphoinositide-specific phospholipase C (PLC) inhibitor U73122 (1 μM; Wako), or GPR40 antagonist GW1100 (10 μM; Merck Millipore) for 20 min prior to the addition of decanoate (C10:0).

Nonaka H., Ohue-Kitano R., Masujima Y., Igarashi M, & Kimura I. (2022). Dietary Medium-Chain Triglyceride Decanoate Affects Glucose Homeostasis Through GPR84-Mediated GLP-1 Secretion in Mice. Frontiers in Nutrition, 9, 848450.

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Culturing and Labeling H1299 Lung Cancer Cells

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Human non-small cell lung carcinoma cell line NCI-H1299 (American Type Culture
Collection, Manassas, VA, USA) was cultured in Roswell Park Memorial Institute (RPMI)
medium (Thermo Fisher Scientific, Waltham, MA, USA) supplemented with 10% fetal bovine
serum (FBS; Funakoshi, Tokyo, Japan) and 1% penicillin streptomycin (Thermo Fisher
Scientific, Waltham, MA, USA) at 37 °C and 5% CO₂. Cells were harvested at 80%
confluence by trypsinization and suspended at 1 × 10⁵ cells per milliliter in
culture medium for cell deposition experiments. The collected cells were incubated in
phosphate-buffered saline with 1 nM calcein-AM (Dojindo Laboratories, Kumamoto, Japan) at
37 °C and 5% CO₂. Trypan blue solution, 0.4% (Thermo Fisher Scientific,
Waltham, MA, USA), was applied to the deposited single cells for live–dead cell
staining.

Onoshima D., Hattori Y., Yukawa H., Ishikawa K., Hori M, & Baba Y. (2018). Cell Deposition Microchip with Micropipette Control over Liquid Interface Motion. Cell Medicine, 10, 2155179017733152.

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