F12 medium

Manufactured by Corning

Sourced in United States

F12 medium is a cell culture medium designed for the growth and maintenance of various cell types. It provides the necessary nutrients and components to support the in vitro cultivation of cells. The medium is formulated to support the specific requirements of the targeted cell lines.

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Close spelling variants of this product

Dulbecco’s modified Eagle’s medium/F12 DMEM/F12 50/50 medium Dulbecco’s Modified Eagle Medium/F12 DMEM/F12 medium Dulbecco’s Modified Eagle Medium/Nutrient Mixture F-12 (DMEM/F12) DMEM/F12 culture medium DMEM/F12 phenol-free medium Dulbecco’s modified Eagle’s medium/F12 (DMEM-F12) Dulbecco’s Modified Eagle Medium/Ham’s Nutrient Mixture F12 (DMEM/F12; DMEM/F12) complete medium

7 protocols using f12 medium

Cell Line Maintenance Conditions

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All cell lines were maintained at 37°C under humidified conditions and 5% CO₂. HEK293 cells were maintained in DMEM medium (Corning, Manassas, VA, USA) supplemented with 10% newborn calf serum (Atlanta Biologicals, Flowery Branch, GA, USA). CHO cells were maintained in 50% F-12 medium (Corning, Manassas, VA, USA) and 50% DMEM medium supplemented with 10% calf serum (Atlanta Biologicals, Flowery Branch, GA, USA).

Neumann B., Chang C.C, & Chang T.Y. (2019). Triton X-100 or octyl glucoside inactivates acyl-CoA:cholesterol acyltransferase 1 by dissociating it from a two-fold dimer to a two-fold monomer. Archives of biochemistry and biophysics, 671, 103-110.

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Cell Culture Protocol for SARS-CoV-2 Research

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Calu3, H1650, African green monkey kidney cell line Vero E6, and HEK293T cells were purchased from the cell bank of the Chinese Academy of Sciences (Shanghai, China). Primary human AT II was purchased from Procell Co. Ltd (Wuhan, China) and cultured with F12 medium (CORNING, NY, USA) supplemented with 10% FBS, and 1% penicillin/streptomycin at 37 °C and 5% CO₂. Prior to experiments, all cell lines were authenticated by genotyping of short tandem repeats using Biowing Biotechnology (Shanghai, China). Additionally, all cell cultures were tested for mycoplasma contamination every 3 months. Human iPSC‐derived lung organoids were generated by Seven Plus Limited Company (Tianjin, China) as previously described.^[⁵⁵^] Further details on cell culture can be found in the Supporting Information.

Liu W., Zhao Y., Fan J., Shen J., Tang H., Tang W., Wu D., Huang W., Ding Y., Qiao P., Lin J., Li Z., Li Q., Cui Q., Liu Y., Chen Y., Pu R., Han X., Yin J., Tan X, & Cao G. (2023). Smoke and Spike: Benzo[a]pyrene Enhances SARS‐CoV‐2 Infection by Boosting NR4A2‐Induced ACE2 and TMPRSS2 Expression. Advanced Science, 10(26), 2300834.

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Knockdown of TSPAN4 in glioma cell lines

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Three glioma cell lines, U251, U87, and T98G, were obtained from the Cell Bank of the Chinese Academy of Sciences (Shanghai, China). Glioma cells were cultured in F12 medium (Corning, NY, USA) mixed with 10% foetal bovine serum (Gibco, MA, USA) in an incubator at 37 °C and 5% CO₂ and used as an in vitro model for analysis. Small interfering RNAs targeting TSPAN4 were designed and synthesized by GenePharma (Shanghai, China) to knock down the expression of TSPAN4. Transfection was performed according to the manufacturer’s instructions, and the cells were inoculated into six-well plates and transfected when they reached 70% confluence. To obtain the siRNA dilution solution, 100 pmol siRNA was diluted with 250 µl Opti-MEM medium, added with 5 µl lipofectamine 2000 to 250 µl Opti-MEM medium, and incubated for 5 min at room temperature. The liquid obtained from the above two steps was mixed and incubated for 20 min at room temperature to obtain a transfection solution (final siRNA concentration of 33 nM) and added to 6-well plates. Subsequently, the medium was changed after 6 h of transfection.
The siRNA target sequences are as follows: TSPAN4-Homo-400; sense (5′-3′): GUGCCAUCAAGGAGAACAATT; and antisense(5′-3′): UUGUUCUCCUUGAUGGCACTT.

Li Y.C., Wu Y., Chen G., Zhu L.Z., Luo X., Nie Q.Q., Zhang L, & Zuo C.J. (2023). Tetraspanins predict the prognosis and characterize the tumor immune microenvironment of glioblastoma. Scientific Reports, 13, 13317.

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Cell Culture Conditions for MCF10A, MCF7, and A549

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MCF10A cells were purchased from the American Type Culture Collection (ATCC) and were cultured in DMEM/F12 (1:1) medium (Gibco) with 5% horse serum (Gibco), 100 μg/ml of human epidermal growth factor (PeproTech), 10 mg/ml of insulin (Sigma), 10 mg/ml of hydrocortisone (Sigma), 0.5 mg/ml of cholera toxin (Sigma), and 1× penicillin-streptomycin (Gibco). MCF7 cells were purchased from ATCC and cultured in EMEM medium (Gibco) with 10% FBS (Gibco), 10 mg/ml of insulin, and 1× penicillin-streptomycin. A549 cells were purchased from ATCC and were cultured in F12 medium (Corning) with 10% FBS and 1× penicillin-streptomycin. All cells were incubated at 37 °C with 5% CO₂.

Zhang J., Tian X.J., Chen Y.J., Wang W., Watkins S, & Xing J. (2018). Pathway crosstalk enables cells to interpret TGF-β duration. NPJ Systems Biology and Applications, 4, 18.

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Lipid Homeostasis Regulation in Cell Lines

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AML12 cells were maintained in F12/DMEM 50/50 medium (Corning, Corning, NY and 10% FBS (Corning, Corning, NY) with supplementation with Insulin-Transferrin-Selenium (Corning, Corning, NY). HepG2 cells were maintained in Eagle’s Minimum Essential Medium (EMEM) with 10% FBS. CHO-K1 cells were maintained in F12 medium (Corning, Corning, NY) and 10% FBS. For Betulin (Sigma) experiments, AML12 cells were pre-treated with 6 μm Betulin for 3 hours before experiment and then changed to medium with 0.5% Methyl-beta-cyclodextrin (MBCD) (Alfa Aesar, Havervill, MA) for additional 6 hours. For siRNA knockdown studies, AML12 cells were plated at 50% confluence one day before transfection. Cells were transfected with 50–80 nM scramble or targeted siRNAs together with lipofectamine 2000 (Life Technologies, Eugene, OR) for 6 hours in reduced serum MEM. AML12 cells were incubated with lipoprotein depleted serum (LPDS) (Alfa Aesar, Havervill, MA) medium for additional 24 hours before harvest. For truncated Srebp2, Srebp2 plasmid was transfected into AML12 cells for 48 hours. For MBCD dose response, AML12 cells were treated with 0%, 0.125%, 0.25% or 0.5% MBCD in 0.5% LPDS medium for 6 hours. For statin treatment cell culture experiments, 6 hours after SiRNA transfection, AML12 cells were treated with 2 μM Mevastatin for additional 16 hours.

Li Z., Votava J.A., Zajac G.J., Nguyen J.N., Jaimes F.B., Ly S.M., Brinkman J.A., De Giorgi M., Kaul S., Green C.L., St. Clair S.L., Belisle S.L., Rios J.M., Nelson D.W., Sorci-Thomas M.G., Lagor W.R., Lamming D.W., Yen C.L, & Parks B.W. (2020). Integrating mouse and human genetic data to move beyond GWAS and identify causal genes in cholesterol metabolism. Cell metabolism, 31(4), 741-754.e5.

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Purification and Expression of ACAT1 Variants

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AC29 cells, an ACAT1-deficent Chinese hamster ovary (CHO) mutant cell line [28 (link), 29 (link)], were maintained in 50% F-12 medium (Corning, Manassas, VA, USA) and 50% DMEM medium supplemented with 10% calf serum (Atlanta Biologicals, Flowery Branch, GA, USA). HEK293 cells were maintained in DMEM medium supplemented with 10% newborn calf serum (Atlanta Biologicals, Flowery Branch, GA, USA). All cell lines were maintained at 37°C under humidified conditions and 5% CO₂.
The expression plasmid HisACAT1/FLAG was constructed as described previously [30 (link)]. In brief, an octapeptide FLAG-tag was inserted at the C-terminus of human ACAT1 cDNA containing a 6-histidine (His)-tag at the N-terminus. This fragment (HisACAT1/FLAG) was ligated into the mammalian expression vector pAG3-Zeo [31 (link)]. The expression plasmid Δ1-65 HisACAT1/FLAG was constructed in a similar manner, by using the HisACAT1 cDNA devoid of the first 65 amino acids [32 (link)]. HisACAT1/FLAG was purified to homogeneity as previously described [12 (link), 30 (link)] from stably expressing HEK293 cells by promptly purifying the enzyme first with HisPur Ni-NTA resin and subsequently with anti-FLAG M2 resin. AC29 cells stably expressing HisACAT1/FLAG or Δ1-65 HisACAT1/FLAG were isolated as previously described [12 (link)].

Neumann B., Chao K., Chang C.C, & Chang T.Y. (2020). Nanodisc scaffold peptide (NSPr) replaces detergent by reconstituting acyl-CoA:cholesterol acyltransferase 1 into peptidiscs. Archives of biochemistry and biophysics, 691, 108518.

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ARPE-19 and MIO-M1 Cell Culture Protocols

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Cell culture: ARPE-19 cells (ATCC, Manassas, VA;) were cultured in Dulbecco's modified Eagle's medium (DMEM) mixture 1:1 Ham's F-12 medium (Corning -Cellgro, Mediatech, Manassas, VA), containing 10% fetal bovine serum (FBS), penicillin G 100 U/mL, streptomycin sulfate 0.1 mg/mL, gentamicin 10 mg/mL, and amphotericin B 2.5 mg/mL. Serum-free medium was used after cells reached monolayer confluence. Human MIO-M1 cells, obtained from the Department of Cell Biology of the University College, London, 63 were grown in Dulbecco's modified medium 1X with high glucose (DMEM+GlutaMAX; Gibco, Carlsbad, CA). Initially, cells were cultured in 10% FBS and penicillin G 100 U/mL, streptomycin sulfate 0.1 mg/mL, but to keep the cells in the non-proliferating phase, the culture media were changed to 2% FBS. Both cultured ARPE-19 and MIO-M1 cells were kept under standard incubating conditions: 37 o C and 5% carbon dioxide.

Moustafa M.T., Ramirez C., Schneider K., Atilano S.R., Limb G.A., Kuppermann B.D, & Kenney M.C. (2017). Protective Effects of Memantine on Hydroquinone-Treated Human Retinal Pigment Epithelium Cells and Human Retinal Müller Cells. Journal of ocular pharmacology and therapeutics : the official journal of the Association for Ocular Pharmacology and Therapeutics, 33(8).

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