Dmem ham s f 12 medium

Manufactured by Harvard Bioscience

Sourced in Germany

DMEM/HAM's F-12 medium is a widely used cell culture medium that supports the growth and maintenance of a variety of cell types. It is a 1:1 mixture of Dulbecco's Modified Eagle's Medium (DMEM) and Ham's F-12 Nutrient Mixture, providing a balanced formulation of essential nutrients, vitamins, and growth factors required for cell proliferation.

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Lab products found in correlation

Fbs superior, by Harvard Bioscience (5 mentions) Penicillin streptomycin, by Harvard Bioscience (5 mentions) Epidermal growth factor (egf), by Merck Group (5 mentions) Amphotericin b, by Merck Group (3 mentions) Penicillin, by Merck Group (3 mentions) Streptomycin, by Merck Group (3 mentions) Hydrocortisone, by Merck Group (3 mentions) Fetal bovine serum (fbs), by Harvard Bioscience (3 mentions) Dmem ham s f12, by Thermo Fisher Scientific (2 mentions) Glutamine, by Harvard Bioscience (2 mentions) Rpmi 1640 medium, by Harvard Bioscience (2 mentions) Cholera toxin, by Merck Group (2 mentions) Insulin transferrin selenium supplement, by Thermo Fisher Scientific (2 mentions) Epidermal growth factor (egf), by Thermo Fisher Scientific (2 mentions) Dmem medium and hanks bss, by GE Healthcare (2 mentions) Collagenase type 2, by Thermo Fisher Scientific (1 mentions) Collagenase, by Thermo Fisher Scientific (1 mentions) Antibiotic solution, by Merck Group (1 mentions) Dmem ham s f12, by Harvard Bioscience (1 mentions) Adenin, by Merck Group (1 mentions)

19 protocols using dmem ham s f 12 medium

Isolation and Culture of Human Chondrocytes

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Human immortalized C28/I2 cells, originating from cells isolated from rib cartilage,21 were cultured in Dulbecco's modified Eagle's medium (DMEM)/HAM's F‐12 medium (Biochrom, Darmstadt, Germany) supplemented with 5 % fetal bovine serum (FBS Superior; Biochrom) and antibiotic‐antimycotic solution (100 U/ml penicillin, 0.1 mg/ml streptomycin, 0.25 µg/ml amphotericin‐B; Sigma‐Aldrich) in a humidified atmosphere at 37 °C and 5 % CO₂.
Human primary chondrocytes were isolated from total knee arthroplasty samples with informed consent and ethical approval by the Ethics Committee of Salzburg (415‐E/1965/4‐2015). All patients were female and all samples were macroscopically graded between 3 and 4 or 4 (Outerbridge classification). Cartilage samples of the whole tibial area and femur condyles were removed from subchondral bone, minced, washed in phosphate‐buffered saline (PBS) and digested overnight in DMEM/HAM's F‐12 supplemented with FBS and 2 mg/ml collagenase Type II (275 units/mg; Thermo Fisher Scientific) at 37 °C on a shaker. After 24 h, the cells were centrifuged, washed twice with PBS, resuspended in medium and seeded as required for the following experiments. For 7‐hydroxy‐3H‐phenoxazin‐3‐one‐10‐oxide sodium salt (Resazurin) assays and Fura‐2 assays, cells were always used at the first passage.

Winklmayr M., Gaisberger M., Kittl M., Fuchs J., Ritter M, & Jakab M. (2019). Dose‐Dependent Cannabidiol‐Induced Elevation of Intracellular Calcium and Apoptosis in Human Articular Chondrocytes. Journal of Orthopaedic Research, 37(12), 2540-2549.

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Culturing C28/I2 and Primary Chondrocytes

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Human immortalized C28/I2 cells (Goldring et al., 1994 (link); Finger et al., 2003 (link)) were cultured in 25 cm² flasks with DMEM/HAM’s F-12 medium (Biochrom, Berlin, Germany) supplemented with 5% fetal bovine serum (FBS Superior, Biochrom) and antibiotic-antimycotic solution (100 U/ml penicillin, 0.1 mg/ml streptomycin, 0.25 μg/ml amphotericin-B; Sigma–Aldrich). C28/I2 cells were kept at 37°C in a humified atmosphere of 5% CO₂ (standard culture conditions). Subcultures were established once a week until passage 25. Primary chondrocytes were isolated from total human knee arthroplasty samples with informed consent and ethical approval by the Ethics Committee of Salzburg (415-E/1965/4-2015) as described in Winklmayr et al. (2019) (link). In brief, cartilage samples were removed from subchondral bone and crashed into small pieces. After washing with phosphate-buffered saline (PBS), they were put on a shaker and digested overnight in DMEM/HAM’s F-12 supplemented with FBS and 2 mg/ml collagenase (Thermo Fisher) at 37°C. After 24 h, cells were (1) centrifuged, (2) washed twice with PBS, (3) resuspended in medium, and (4) seeded as required for the following experiments. Primary chondrocytes were used up to passage 2.

Kittl M., Winklmayr M., Helm K., Lettner J., Gaisberger M., Ritter M, & Jakab M. (2020). Acid- and Volume-Sensitive Chloride Currents in Human Chondrocytes. Frontiers in Cell and Developmental Biology, 8, 583131.

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Characterization of MLH1 Null Tumor Cell Lines

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Two MLH1^−/− tumor cell lines 328 and A7450 T1 M1 were established and characterized in our lab.^{25 (link),26 (link)} Cell culture was performed in DMEM/Ham’s F12 medium, supplemented with 10% FCS (fetal calf serum), 6 mM Glutamine, and antibiotics (all from Biochrom, Berlin, Germany). Treatment was done with selected CDKIs at doses corresponding to IC₃₀ values: (I) abemaciclib (= abema): 1 µM; (II) dinaciclib (= dina): 100 nM; (III) THZ-1: 0.83 µM. Doses were validated before via dose response curve analyses.

Salewski I., Henne J., Engster L., Krone P., Schneider B., Redwanz C., Lemcke H., Henze L., Junghanss C, & Maletzki C. (2022). CDK4/6 blockade provides an alternative approach for treatment of mismatch-repair deficient tumors. Oncoimmunology, 11(1), 2094583.

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Kinase Inhibitor Profiling using Cell Lines

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The cell lysate mixture (cell line mix) used to profile all kinase inhibitors in this study was generated from K-562, COLO 205 and MV-4-11 cells grown in RPMI1640 medium (Biochrom GmbH), SK-N-BE(2) cultured in DMEM/HAM’s F-12 medium (Biochrom GmbH). All were supplemented with 10% FBS (Biochrom GmbH) and 1% antibiotic solution (Sigma). Cell lines were authenticated by multiplex human cell line authentication test (MCA) and tested internally for mycoplasma contamination. For MET-inhibitor profiling, Caki-1 cells were cultured in IMDM (Biorad) with 10% FBS. For EGFR-inhibitor profiling, BT-474 cells were grown in DMEM/HAM’s F-12 supplemented with 15% FBS (Biochrom). Kinase inhibitor affinity matrices (Kinobeads) were synthesized in house as published(14 (link)). Omipalisib with a linker was internally synthesized. Small molecule kinase and other inhibitors were purchased from Selleckchem, MedChemExpress, Active Biochem, Abmole, Merck or LC Labs.

Klaeger S., Heinzlmeir S., Wilhelm M., Polzer H., Vick B., Koenig P.A., Reinecke M., Ruprecht B., Petzoldt S., Meng C., Zecha J., Reiter K., Qiao H., Helm D., Koch H., Schoof M., Canevari G., Casale E., Re Depaolini S., Feuchtinger A., Wu Z., Schmidt T., Rueckert L., Becker W., Huenges J., Garz A.K., Gohlke B.O., Zolg D.P., Kayser G., Vooder T., Preissner R., Hahne H., Tonisson N., Kramer K., Goetze K., Bassermann F., Schlegl J., Ehrlich H.C., Aiche S., Walch A., Greif P.A., Schneider S., Felder E.R., Ruland J., Medard G., Jeremias I., Spiekermann K, & Kuster B. (2017). The target landscape of clinical kinase drugs. Science (New York, N.Y.), 358(6367), eaan4368.

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Generation of Vinculin and E-cadherin Deficient Keratinocytes

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Vinculin and E-cadherin-deficient cells were generated from epidermal, murine keratinocytes as described before (Michels et al., 2009 (link); Mertz et al., 2013 (link); Rubsam et al., 2017 (link)). All cells were cultivated under sterile conditions at 32°C and 5% (vol/vol) CO₂ in low calcium (<0.5 mM) containing DMEM/Ham´s F12 medium (Biochrom, Berlin, Germany) supplemented with 10% FCS Gold (chelex treated) (PAA laboratories, Cölbe, Germany), 1% glutamine (Biochrom, Berlin, Germany), 1% penicillin/streptomycin (Biochrom, Berlin, Germany), 0.18 mM adenin (Sigma, St. Louis, MO), 0.5 µg/ml hydrocortisone (Sigma), 5 µg/ml insulin (Sigma), 10 ng/ml EGF (Sigma), 0.05 µg/µl choleratoxin (Sigma), and 5 µg/ml vitamin C (Sigma).

Noethel B., Ramms L., Dreissen G., Hoffmann M., Springer R., Rübsam M., Ziegler W.H., Niessen C.M., Merkel R, & Hoffmann B. (2018). Transition of responsive mechanosensitive elements from focal adhesions to adherens junctions on epithelial differentiation. Molecular Biology of the Cell, 29(19), 2317-2325.

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Prostate Carcinoma Cell Lines Protocol

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Two human and one canine prostate carcinoma cell line were used. The human PC-3 cell line (24 (link)) (DSMZ Cat# ACC-465, RRID:CVCL_0035) was cultivated in DMEM/Ham's F-12 medium (Biochrom GmbH, Berlin, Germany). The human LNCaP cell line (25 (link)) (DSMZ Cat# ACC-256, RRID:CVCL_0395) was grown in RPMI 1640 medium (Biochrom GmbH). The canine cell line TihoDProAdcarc1258 (26 (link)) (CT1258, RRID:CVCL_W737) was established by our group and cultivated in medium 199 (Live Technologies GmbH, Darmstadt, Germany). All media were supplemented with 10% heat-inactivated fetal bovine serum (FBS Superior, Biochrom GmbH) and 2% penicillin/streptomycin (Biochrom GmbH). All cells were cultivated at 37°C in a humidified atmosphere of 5% CO₂.

Schille J.T., Nolte I., Beck J., Jilani D., Roolf C., Pews-Davtyan A., Rolfs A., Henze L., Beller M., Brenig B., Junghanss C., Schütz E, & Murua Escobar H. (2021). PDA Indolylmaleimides Induce Anti-Tumor Effects in Prostate Carcinoma Cell Lines Through Mitotic Death. Frontiers in Veterinary Science, 7, 558135.

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Osteoblast Cytocompatibility of Porous Titanium Implants

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Primary human osteoblast-like cells from four donors (2 males and 2 females, aged 60–85 years, cells isolated from the femoral head, IRB approval: EA4/035/14) in passage 2 were pooled. The patients gave written informed consent. Cells were cultured in DMEM/Ham's F-12 medium (Biochrom, Berlin, Germany) with 10% fetal bovine serum (FBS; Biochrom, Berlin, Germany), 1% penicillin/streptomycin (P/S; Biochrom, Berlin, Germany) and freshly supplemented 0.05 mM l-ascorbic acid (L-Asc) and 0.05 mM β-glycerol phosphate (β-Gly) (both Sigma-Aldrich/Merck, Darmstadt, Germany), henceforth called growth medium (GM).
The following groups were analyzed: 1) cells only (Medium), 2) titanium K-wire (CTRL), 3) porous K-wire (Porous), and 4) porous K-wire loaded with gentamicin (Genta). The porous K-wires of group 3 and 4 were coated with silver nanoparticles. For the cytocompatibility test, a toxic control was used: 5) 10% 2-hydroxyethyl methacrylate (HEMA; Positive). The experiments were done in triplicate and repeated once, resulting in n = 6 per group. For the experiments, 1.4 mm K-wire pieces (7 mm length) were transferred to inserts (10 mm diameter and polycarbonate membrane with 8 μm pores; Thermo Scientific/Nunc, Schwerte, Germany) and placed in the culture wells.

Ständert V., Borcherding K., Bormann N., Schmidmaier G., Grunwald I, & Wildemann B. (2021). Antibiotic-loaded amphora-shaped pores on a titanium implant surface enhance osteointegration and prevent infections. Bioactive Materials, 6(8), 2331-2345.

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Cell-SELEX on Breast Cancer Cell Lines

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Human breast cancer MDA-MB-231 (ATCC HTB-26) and non-tumorigenic epithelial human breast MCF-10-2A (ATCC CRL-10781) cell lines were used for the cell-SELEX as target cells and negative control, respectively. Hs 578T (ATCC HTB-126), MDA-MB-468 (ATCC HTB-132), MDA-MB-453 (ATCC HTB-131), MCF-7 (ATCC HTB-22) and MDA-MB-157 (ATCC HTB-24) cell lines were also used to confirm the aptamers binding specificity. MDA-MB-231, Hs 578T, MDA-MB-468, MDA-MB-453, MCF-7 and MDA-MB-157 cells were cultured in Dulbecco’s Modified Eagle Medium (DMEM) [Biochrom] supplemented with 10% Fetal Bovine Serum (FBS) [Biochrom] and 1% penicillin–streptomycin [Biochrom] on tissue culture treated flasks. MCF-10-2A cells were cultured in a 1:1 mixture of DMEM:Ham’s F12 medium [Biochrom] supplemented with 20 ng/ml Epidermal Growth Factor (EGF) [Sigma], 100 ng/ml cholera toxin [Sigma], 0.01 mg/ml insulin [Biochrom], 500 ng/ml hydrocortisone [Sigma], 5% horse serum [Biochrom] and 1% penicillin–streptomycin. Cells were propagated at 37 °C, 5% CO₂ and 95% relative humidity. The cells were washed using phosphate-buffered saline (PBS) 1X pH 7.4 and were detached using Trypsin/EDTA solution [Biochrom]. Cells were counted in a hemocytometer using the trypan blue exclusion test.

Ferreira D., Barbosa J., Sousa D.A., Silva C., Melo L.D., Avci-Adali M., Wendel H.P, & Rodrigues L.R. (2021). Selection of aptamers against triple negative breast cancer cells using high throughput sequencing. Scientific Reports, 11, 8614.

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Osteogenic Differentiation of MSCs

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MSCs were isolated from cancellous bone of femoral heads obtained from patients undergoing total hip replacement surgery. Isolation was performed as mentioned previously [38 , 39 (link)]. In brief, cancellous bone fragments were washed thoroughly with PBS and PBS wash solution centrifuged at 400 x g for 5 min. The cell pallet was then collected in Dulbecco’s Medium Essential Medium (DMEM)/Ham’s F-12 medium (Biochrom, Berlin, Germany) supplemented with 20% (v/v) fetal bovine serum (FBS) (Sigma, Taufkirchen, Germany) and 1% (v/v) Penicillin/Streptomycin (Biochrom, Berlin, Germany) and seeded at a density of 2x10⁶ cells/cm² in T175 flasks coated with collagen type I (Corning, Bedford, MA, USA). After splitting cells culturing was continued using 10% (v/v) FBS. In passage 2 osteogenic differentiation of MSCs was induced by incubation in osteogenic differentiation medium (ODM) for a minimum of two weeks consisting of DMEM/Ham’s F-12, 10% (v/v) FBS, 1% (v/v) Pen/Strep, 10 mM β-glycerophosphate (Sigma-Aldrich, Darmstadt, Germany), 0.1 μM dexamethasone (Sigma-Aldrich, Darmstadt, Germany) and 50 μM ascorbic acid (Sigma-Aldrich, Darmstadt, Germany). Medium was changed three times a week. Experiments with MSCs were conducted with cells in passage no. 4 and 5.

Rasch A., Naujokat H., Wang F., Seekamp A., Fuchs S, & Klüter T. (2019). Evaluation of bone allograft processing methods: Impact on decellularization efficacy, biocompatibility and mesenchymal stem cell functionality. PLoS ONE, 14(6), e0218404.

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Detailed Cell Culture Protocol

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DMEM/Ham’s F12 medium was purchased from Biochrom AG (Berlin, Germany), DMEM medium and Hank´s BSS from PAA Laboratories (Coelbe, Germany), insulin-transferrin-selenium supplement from Gibco (Karlsruhe, Germany), fetal calf serum (FCS) from PAN Biotech (Aidenbach, Germany), triiodothyronine from Fluka (Buchs, Switzerland), hydrocortisone and lysophosphatidic acid (LPA) from Sigma-Aldrich (Munich, Germany), epidermal growth factor from PeproTech (Hamburg, Germany), Y27632, (+)-(R)-trans-4-(1-aminoethyl)-N-(4-pyridyl) cyclohexanecarboxamide dihydrochloride from Calbiochem, H1152 (S)-(+)-2-methyl-1-[(4-methyl-5-isoquinolinyl)sulfonyl]-homopiperazine from Alexis Biochemicals (Grünberg, Germany), and EHT1864 (5-(5-(7-(Trifluoromethyl)quinolin-4-ylthio)pentyloxy)-2-(morpholinomethyl)-4H-pyran-4-one dihydrochloride) from Sigma-Aldrich.

Giehl K., Keller C., Muehlich S, & Goppelt-Struebe M. (2015). Actin-Mediated Gene Expression Depends on RhoA and Rac1 Signaling in Proximal Tubular Epithelial Cells. PLoS ONE, 10(3), e0121589.

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