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Freestyle f 17 medium

Manufactured by Thermo Fisher Scientific
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The Freestyle F-17 medium is a laboratory equipment product designed for cell culture applications. It serves as a growth medium for the cultivation of various cell types. The product's core function is to provide the necessary nutrients and support for the proliferation and maintenance of cultured cells.

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Lab products found in correlation

Glutamax, by Thermo Fisher Scientific (9 mentions) Hek293f cell, by Thermo Fisher Scientific (6 mentions) Transit boost, by Mirus Bio (5 mentions) Pluronic f 68, by Thermo Fisher Scientific (2 mentions) Geneticin, by Thermo Fisher Scientific (2 mentions) L glutamine, by Thermo Fisher Scientific (2 mentions) Linear pei 25 kda, by Polysciences (1 mentions) Poloxamer 188, by Corning (1 mentions) Pluronic f68, by AppliChem (1 mentions) Hispur ni nta resin, by Thermo Fisher Scientific (1 mentions) Superdex 200, by GE Healthcare (1 mentions) Histrap excel, by GE Healthcare (1 mentions) Ex cell 302 serum free medium, by Merck Group (1 mentions) Hek293t cell, by American Type Culture Collection (1 mentions) Kolliphor p188, by Merck Group (1 mentions) Pei max reagent, by Polysciences (1 mentions) Insect xpress medium, by Lonza (1 mentions)

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Freestyle medium (54 citations)

17 protocols using freestyle f 17 medium

1

Transient Expression of A2M Variants

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Example 9

A2M variants were expressed in HEK293F cells (Gibco) by transient transfection of each construct in suspension cells. Cells were grown to a density of 550,000 cells/mL in a Erlenmeyer cell culture flask containing 20 mL of FreeStyle F17 medium (Invitrogen) containing 1× GlutaMax (Gibco) on a rotator at a speed of 125 rpm inside a 37° C. incubator containing an 8% CO2/air mixture. Cells were transfected by mixing 20 μg of plasmid DNA of each construct (wild-type or variant) in a 1:2 (w/v) ratio with TransIT Pro plus 10 μL TransIT Boost (Mirus) 15 minutes before addition to media. Cells were maintained in the same conditions for three days after transfection before the media containing secreted recombinant protein was removed for protein purification (FIG. 3).

US10889631B2. Therapeutic variant alpha-2-macroglobulin compositions (2021-01-12). CYTONICS CORPORATION [US]. Inventors: Lewis Hanna [US], John David Laughlin [US], Shawn Robert Browning [US].
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2

Transient Transfection of A2M Variants in HEK293F Cells

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Example 9

A2M variants were expressed in HEK293F cells (Gibco) by transient transfection of each construct in suspension cells. Cells were grown to a density of 550,000 cells/mL in a Erlenmeyer cell culture flask containing 20 mL of FreeStyle F17 medium (Invitrogen) containing 1× GlutaMax (Gibco) on a rotator at a speed of 125 rpm inside a 37° C. incubator containing an 8% CO2/air mixture. Cells were transfected by mixing 20 μg of plasmid DNA of each construct (wild-type or variant) in a 1:2 (w/v) ratio with TransIT Pro plus 10 μL TransIT Boost (Mirus) 15 minutes before addition to media. Cells were maintained in the same conditions for three days after transfection before the media containing secreted recombinant protein was removed for protein purification (FIG. 3).

US10940189B2. Systems, compositions, and methods for transplantation (2021-03-09). Cytonics Corporation [US]. Inventors: Lewis Hanna [US], John David Laughlin [US], Shawn Robert Browning [US].
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3

Recombinant A2M Variant Expression in HEK293F

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Example 9

A2M variants were expressed in HEK293F cells (Gibco) by transient transfection of each construct in suspension cells. Cells were grown to a density of 550,000 cells/mL in a Erlenmeyer cell culture flask containing 20 mL of FreeStyle F17 medium (Invitrogen) containing 1× GlutaMax (Gibco) on a rotator at a speed of 125 rpm inside a 37° C. incubator containing an 8% CO2/air mixture. Cells were transfected by mixing 20 μg of plasmid DNA of each construct (wild-type or variant) in a 1:2 (w/v) ratio with TransIT Pro plus 10 μL TransIT Boost (Mirus) 15 minutes before addition to media. Cells were maintained in the same conditions for three days after transfection before the media containing secreted recombinant protein was removed for protein purification (FIG. 3).

US11040092B2. Systems, compositions, and methods for transplantation (2021-06-22). Cytonics Corporation [US]. Inventors: Lewis Hanna [US], John David Laughlin [US], Shawn Robert Browning [US].
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4

Recombinant A2M Variant Expression

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Example 9

A2M variants were expressed in HEK293F cells (Gibco) by transient transfection of each construct in suspension cells. Cells were grown to a density of 550,000 cells/mL in a Erlenmeyer cell culture flask containing 20 mL of FreeStyle F17 medium (Invitrogen) containing 1× GlutaMax (Gibco) on a rotator at a speed of 125 rpm inside a 37° C. incubator containing an 8% CO2/air mixture. Cells were transfected by mixing 20 μg of plasmid DNA of each construct (wild-type or variant) in a 1:2 (w/v) ratio with TransIT Pro plus 10 μL TransIT Boost (Minis) 15 minutes before addition to media. Cells were maintained in the same conditions for three days after transfection before the media containing secreted recombinant protein was removed for protein purification (FIG. 3).

US10265388B2. Systems, compositions, and methods for transplantation (2019-04-23). CYTONICS CORPORATION [US]. Inventors: Lewis Hanna [US], John David Laughlin [US], Shawn Robert Browning [US].
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5

Transient Expression of A2M Variants

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Example 9

A2M variants were expressed in HEK293F cells (Gibco) by transient transfection of each construct in suspension cells. Cells were grown to a density of 550,000 cells/mL in a Erlenmeyer cell culture flask containing 20 mL of FreeStyle F17 medium (Invitrogen) containing 1× GlutaMax (Gibco) on a rotator at a speed of 125 rpm inside a 37° C. incubator containing an 8% CO2/air mixture. Cells were transfected by mixing 20 μg of plasmid DNA of each construct (wild-type or variant) in a 1:2 (w/v) ratio with TransIT Pro plus 10 μL TransIT Boost (Mirus) 15 minutes before addition to media. Cells were maintained in the same conditions for three days after transfection before the media containing secreted recombinant protein was removed for protein purification (FIG. 3).

US11634475B2. Therapeutic variant alpha-2-macroglobulin compositions (2023-04-25). CYTONICS CORPORATION [US]. Inventors: Lewis Hanna [US], John David Laughlin [US], Shawn Robert Browning [US].
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6

Transient Expression of A2M Variants

Check if the same lab product or an alternative is used in the 5 most similar protocols

Example 9

A2M variants were expressed in HEK293F cells (Gibco) by transient transfection of each construct in suspension cells. Cells were grown to a density of 550,000 cells/mL in a Erlenmeyer cell culture flask containing 20 mL of FreeStyle F17 medium (Invitrogen) containing 1× GlutaMax (Gibco) on a rotator at a speed of 125 rpm inside a 37° C. incubator containing an 8% CO2/air mixture. Cells were transfected by mixing 20 mg of plasmid DNA of each construct (wild-type or variant) in a 1:2 (w/v) ratio with TransIT Pro plus 10 μL TransIT Boost (Mirus) 15 minutes before addition to media. Cells were maintained in the same conditions for three days after transfection before the media containing secreted recombinant protein was removed for protein purification (FIG. 3).

US10400028B2. Therapeutic variant alpha-2-macroglobulin compositions (2019-09-03). CYTONICS CORPORATION [US]. Inventors: Lewis Hanna [US], John David Laughlin [US], Shawn Robert Browning [US].
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7

Transient Transfection of CHO and HEK Cells

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β-Klotho and Fc-FGF21 mutant constructs were transiently transfected into CHO-EBNA1 (clone 3E7) and HEK 293-EBNA1 (clone 6E) cells, respectively, as previously described59 (link),60 (link). Briefly, CHO-EBNA1 cells were maintained in a chemically-defined CHO cell culture medium. HEK293-EBNA1 cells were maintained in Freestyle F17 medium (Invitrogen) supplemented with 6 mM GlutaMax (Invitrogen), 0.1% (w/v) poloxamer 188, and 25 µg/mL G418 (Corning). Cells were cultured in suspension format using shaker flasks on an orbital shaking platform rotating at 110 rpm in a humidified incubator with 5% CO2 at 37 °C. Cells were transfected during the exponential growth phase at 1 and 0.5 µg DNA/mL culture for CHO-EBNA1 and HEK293-EBNA1 cells, respectively, using linear PEI 25 kDa (Polysciences Inc.). Conditioned medium was harvested 6 and 7 days post-transfection for HEK293-EBNA1 and CHO-EBNA1 cells, respectively, by centrifuging cells at 4,000 rpm for 30 minutes and filtering the supernatant with a 0.2 µm PES membrane.
Shi S.Y., Lu Y.W., Richardson J., Min X., Weiszmann J., Richards W.G., Wang Z., Zhang Z., Zhang J, & Li Y. (2018). A systematic dissection of sequence elements determining β-Klotho and FGF interaction and signaling. Scientific Reports, 8, 11045.
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8

Transient Protein Production in HEK293-6E Cells

Cited 1 time
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HEK293-6E-MGAT1-k.o. cells were cultivated in FreeStyle F17 medium (ThermoFisher), supplemented with 7.5 mM L-glutamine and 0.1% Pluronic F-68 (AppliChem, Germany), in the presence of 25 μg/mL geneticin. Transient production was performed as described in [45 , 46 (link)]. The supernatants were harvested at time point 168 h by centrifugation for 30 min at 3000 g and prepared for purification.
Büssow K., Themann P., Luu S., Pentrowski P., Harting C., Majewski M., Vollmer V., Köster M., Grashoff M., Zawatzky R., Van den Heuvel J., Kröger A, & Böldicke T. (2019). ER intrabody-mediated inhibition of interferon α secretion by mouse macrophages and dendritic cells. PLoS ONE, 14(4), e0215062.
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9

Engineered hIL-2 Fusion Protein Production

Cited 1 time
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Mutations in the targeted segments of hIL-2 were introduced by site-directed mutagenesis on the template plasmid pCMX containing wt hIL-2 gene, using suitable mutagenic oligonucleotides (CIGB, Cuba). Mutations and correctness of the rest of the gene were confirmed by DNA sequencing (Microsynth-Seqlab, Germany). Modified IL-2 genes were re-cloned into pCSE-2.6 hIgG1 Fc38 (link) through BssHII/NotI restriction sites, and sequenced again. HEK-293T cells adapted to grow in suspension in Freestyle F-17 medium (ThermoFisher, USA) were transiently transfected with the resulting genetic constructs, using linear polyethyleneimine (PEI) as the transfecting agent according to established procedures38 (link). Fusion proteins were purified from cell culture supernatants by protein A affinity chromatography.
Rojas G., Relova-Hernández E., Pérez-Riverón A., Castro-Martínez C., Diaz-Bravo O., Infante Y.C., Gómez T., Solozábal J., DíazBravo A.B., Schubert M., Becker M., Pérez-Massón B., Pérez-Martínez D., Alvarez-Arzola R., Guirola O., Chinea G., Graca L., Dübel S., León K, & Carmenate T. (2023). Molecular reshaping of phage-displayed Interleukin-2 at beta chain receptor interface to obtain potent super-agonists with improved developability profiles. Communications Biology, 6, 828.
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10

Efficient HDAC6 Variant Expression

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All HDAC6 variants were expressed using HEK-293/T17 cells following transient transfection mediated by linear polyethylene imine (PEI) (Polysciences Inc., Warrington, PA, USA). To this end, the suspension culture of HEK-293/T17 cells was grown in 2 L Erlenmeyer flasks in Free Style F17 medium (Thermo Fisher Scientific) supplemented by 0.1% Pluronic F-68 (Invitrogen) and 2 mM L-glutamine at 110 rpm under a humidified 5% CO2 atmosphere at 37 °C. For large-scale expression, 0.7 mg of an expression plasmid was diluted in 17.5 ml of PBS, to which 2.1 ml of 1 mg/ml PEI was added. The mixture was vortexed briefly, incubated for 10 min at room temperature, and then added to 350 ml of cells at a concentration of 4 × 106 cell/ml. Four hours post-transfection, the cell suspension was diluted by an equal volume of ExCell serum-free medium. Cells were harvested 72 h post transfection by centrifugation at 500 × g for 5 min, then the cell pellet was frozen in liquid nitrogen and stored at −80 °C until further use.
Skultetyova L., Ustinova K., Kutil Z., Novakova Z., Pavlicek J., Mikesova J., Trapl D., Baranova P., Havlinova B., Hubalek M., Lansky Z, & Barinka C. (2017). Human histone deacetylase 6 shows strong preference for tubulin dimers over assembled microtubules. Scientific Reports, 7, 11547.
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