The NCH421k, NCH441 and NCH644 glioblastoma sphere cell lines were kindly provided by Professor Christel Herold-Mende from the Department of Neurosurgery at Heidelberg University (Heidelberg, Germany) and were cultivated in serum-free medium, which consisted of Dulbecco's modified Eagle's medium (DMEM)/F-12 medium (Sigma-Aldrich, St. Louis, MO, USA) containing 20% BIT serum-free supplement (STEMCELL Technologies, Inc., Vancouver, BC, Canada), 20 ng/ml basic fibroblast growth factor (provitro GmbH, Berlin, Germany) and 20 ng/ml epidermal growth factor (provitro GmbH). To induce differentiation, glioblastoma spheres were grown in DMEM containing 10% fetal calf serum (Biochrom, Ltd., Cambridge, UK).
Bit serum free supplement
Manufactured by STEMCELL
Sourced in United States
BIT is a serum-free supplement for cell culture. It provides essential components to support cell growth and proliferation.
Automatically generated - may contain errors
Lab products found in correlation
Fetal calf serum (fcs), by Harvard Bioscience (1 mentions)
Dmem f12 medium, by Merck Group (1 mentions)
Basic fibroblast growth factor (bfgf), by Provitro (1 mentions)
Epidermal growth factor (egf), by Provitro (1 mentions)
Human recombinant epidermal growth factor, by STEMCELL (1 mentions)
Human basic fibroblast growth factor, by STEMCELL (1 mentions)
Epidermal growth factor (egf), by Thermo Fisher Scientific (1 mentions)
Basic fibroblast growth factor (bfgf), by Thermo Fisher Scientific (1 mentions)
3 protocols using bit serum free supplement
1
Glioblastoma Sphere Cell Line Protocol
2
Glioblastoma Spheres Generation Protocol
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Gliomaspheres were generated in analogy to well established standard protocols.37 , 39 , 40 , 41 Briefly, glioblastoma cells were transferred from T75 to T25 flasks (to facilitate cell‐cell contacts) with serum‐free media supplemented with growth factors – as typically used for a spheric phenotype15 , 36 , 42 : DMEM/Nutrient Mixture F‐12 medium (DMEM, Gibco, Life Technologies, Paisley, UK) supplemented with 20% BIT‐serum free supplement (bovine serum albumin, insulin, transferrin), human recombinant epidermal growth factor and human basic fibroblast growth factor at 20 ng/mL each (all STEMCELL Technologies, Vancouver, BC, Canada). For passaging and plating, spheres were transferred into conical tubes, centrifuged (200 g, 5 minutes), resuspended in serum‐free medium, dissociated into single‐cell suspensions using trituration (60‐70 times using a Pasteur pipette) and re‐plated. BIT as well as growth factors were added freshly to maintain the concentration after each passage. Tumor spheres were observed under a light microscope every day.
3
Glioblastoma Cell Culture Protocols
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The glioblastoma cell lines U87 and U251 were originally obtained from American Type Culture Collection (ATCC, Rockville, MD, USA). They were cultured in Dulbecco's modified Eagle's medium (DMEM) (Hyclone, Logan, UT, USA) containing 10% (v/v) fetal bovine serum (FBS) (Hyclone, Logan, UT, USA) at 37 °C in a humidified incubator with 5% CO 2 as previously described [24] .
The primary glioblastoma stem-like cell lines, NCH-421k and NCH-644, were obtained from the Department of Neurosurgery at Heidelberg University. They were cultured in serum-free DMEM/F12 medium (Hyclone, Logan, UT, USA) comprising 20 ng/ml epidermal growth factor (EGF) (Peprotech, Rocky Hill, NJ, USA), 20 ng/ml basic fibroblast growth factor (bFGF) (Peprotech, Rocky Hill, NJ, USA) and 20% BIT serum-free supplement (STEMCELL Technologies Inc., Vancouver, BC, Canada).
The primary glioblastoma stem-like cell lines, NCH-421k and NCH-644, were obtained from the Department of Neurosurgery at Heidelberg University. They were cultured in serum-free DMEM/F12 medium (Hyclone, Logan, UT, USA) comprising 20 ng/ml epidermal growth factor (EGF) (Peprotech, Rocky Hill, NJ, USA), 20 ng/ml basic fibroblast growth factor (bFGF) (Peprotech, Rocky Hill, NJ, USA) and 20% BIT serum-free supplement (STEMCELL Technologies Inc., Vancouver, BC, Canada).
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