Drosophila Schneider’s S2 cells (Invitrogen, UK) were cultured in Hyclone SFX-Insect Medium (GE Healthcare, UK) supplemented with 10% (v/v) fetal bovine serum (FBS) (Gibco, UK), 100 U/mL penicillin and 100 μg/mL streptomycin. Cells were incubated at 25 oC and passaged every 3–4 days, when they reached a confluence of ~1 x 107 cells/mL. Human embryonic kidney (HEK 293T) cells were routinely cultured in GlutaMAX Dulbecco’s Modified Eagle Medium (Gibco) supplemented with 10% (v/v) FBS (Gibco), 100 U/mL penicillin and 100 μg/mL streptomycin. Cells were incubated at 37 oC, 5% CO2 and passaged every 3–4 days, when they reached a confluence of ~80%. Cells were detached by incubation for 3–5 min at 37 oC in 5 mM EDTA- phosphate-buffered saline (PBS).
Hyclone sfx insect medium
Manufactured by GE Healthcare
Sourced in United Kingdom, United States
HyClone SFX-insect medium is a serum-free, protein-free, chemically-defined medium developed for the cultivation of insect cells. It is designed to support the growth and production of recombinant proteins in insect cell expression systems.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
Bac to bac baculovirus expression system, by Thermo Fisher Scientific (2 mentions)
Glutamax dulbeccos modified eagle medium, by Thermo Fisher Scientific (1 mentions)
Lucifer yellow, by Merck Group (1 mentions)
L glutamine, by GE Healthcare (1 mentions)
Streptomycin, by Thermo Fisher Scientific (1 mentions)
Penicillin, by Thermo Fisher Scientific (1 mentions)
5 protocols using hyclone sfx insect medium
1
Culturing Drosophila S2 and HEK 293T Cells
2
Insect Cell-based Transporter Assays
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Spodoptera frugiperda (Sf9) insect cells were obtained
from the American Type Culture
Collection (ATCC, USA). The cells were cultured in HyClone SFX Insect
medium (GE Healthcare, USA) supplemented with the fetal bovine serum
(Gibco). Lucifer yellow (LY, Sigma-Aldrich, USA), 5(6)-carboxy-2′,7′-dichlorofluorescein
(CDCF, Sigma-Aldrich, USA), and N-methyl-quinidine
(NMQ, Solvo Biotechnology, Hungary) were used as substrate probes.
The 44 tested inhibitors were provided by the University of Pittsburgh
Chemical Methodologies and Library Development Center (UPCMLD, USA)
and were described previously.29 (link),30 (link) The remainder of the
chemicals were purchased from Sigma-Aldrich (USA) unless stated otherwise.
from the American Type Culture
Collection (ATCC, USA). The cells were cultured in HyClone SFX Insect
medium (GE Healthcare, USA) supplemented with the fetal bovine serum
(Gibco). Lucifer yellow (LY, Sigma-Aldrich, USA), 5(6)-carboxy-2′,7′-dichlorofluorescein
(CDCF, Sigma-Aldrich, USA), and N-methyl-quinidine
(NMQ, Solvo Biotechnology, Hungary) were used as substrate probes.
The 44 tested inhibitors were provided by the University of Pittsburgh
Chemical Methodologies and Library Development Center (UPCMLD, USA)
and were described previously.29 (link),30 (link) The remainder of the
chemicals were purchased from Sigma-Aldrich (USA) unless stated otherwise.
3
Baculovirus Expression in Sf9 Cells
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Spodoptera frugiperda (Sf9) cells were cultured in HyClone SFX-insect medium (GE Healthcare), usually as liquid cultures on a horizontal shaker at 120 rpm and 27°C. The Bac-to-Bac Baculovirus Expression System (Invitrogen) was used to generate bacmids and baculoviruses.
4
Baculovirus-Mediated Protein Expression in Sf9 Cells
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Sf9 insect cells were grown in HyClone SFX-insect medium (GE Healthcare), without supplements and incubated at 27°C either as a monolayer culture or in suspension. Bacmids and baculoviruses were generated using the Bac-to-Bac Baculovirus Expression System (Invitrogen). To express a protein of interest, 2 × 106 cells/ml were infected at an estimated MOI of 1, and incubated for 48 h at 27°C on a horizontal shaker at 200 rpm. Then, the cells were harvested by centrifugation at 475g for 10 min.
5
Cultivation of E. coli and S2 cells
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Transformation-competent E. coli, strains Top10 (ThermoFisher) and BL21 (New England BioLabs), were grown on standard LB+agar with appropriate antibiotic supplement. S2 cells were grown in either HyClone SFX-Insect medium or SFM4-Insect medium plus L-Glutamine (GE Healthcare Life Sciences) supplemented with 100 units/ml penicillin and 100 μg/ml streptomycin (ThermoFisher).
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