Normal human astrocytes (NHA) cell line was purchased from Gibco (N7805100) and cultured in human astrocytes growth medium (Cell Applications). Human neuroblastoma (SH-SY5Y) cell line was obtained from ATCC (CRL-2266) and cultured in DMEM/F12 medium (Gibco). Human glioblastoma (U118) cell line was purchased from ATCC (HTB-15) maintained in Dulbecco's Modified Eagle's Medium (Gibco). In addition, human neural progenitor cells (hNPCs; ReNcell VM) (Takahashi and Yamanaka 2006 (link); Donato et al. 2007 (link)) were commercially purchased from Sigma-Aldrich (SCC008). The cells were grown on 20 μg/mL laminin (Merck)-coated culture plates containing ReNCell NSC maintenance medium (Merck) supplemented with 20 ng/mL of bFGF and EGF (Merck). All culture media were supplemented with 10% fetal bovine serum (FBS) and 1× antibiotic-antimycotic (Gibco) at 37°C with 5% CO2. Cells were passaged when the confluence reached 80% of the culture plate every 3 d. Briefly, cells were rinsed with PBS and then incubated in Accutase (Millipore) for 3–5 min until the cell detached. We used the culture medium to inhibit enzymatic reaction and centrifuged the suspension at 500g for 5 min. We then resuspended the cell pellet in fresh medium and counted the cell number.
Scc008
Manufactured by Merck Group
Sourced in United States
The SCC008 is a versatile laboratory equipment designed for general scientific applications. It functions as a high-performance centrifuge, capable of separating various samples and materials based on their density and size. The SCC008 provides consistent and reliable results, making it a valuable tool for researchers and scientists working in a wide range of disciplines.
Automatically generated - may contain errors
Lab products found in correlation
Rencell vm, by Merck Group (2 mentions)
N7805100, by Thermo Fisher Scientific (1 mentions)
Htb 15, by American Type Culture Collection (1 mentions)
Rencell nsc maintenance medium, by Merck Group (1 mentions)
Sh sy5y, by American Type Culture Collection (1 mentions)
Accutase, by Merck Group (1 mentions)
Antibiotic antimycotic, by Thermo Fisher Scientific (1 mentions)
Dmem f12 medium, by Thermo Fisher Scientific (1 mentions)
Dulbecco s modified eagle s medium, by Thermo Fisher Scientific (1 mentions)
Laminin, by Merck Group (1 mentions)
Heparin sulfate, by Merck Group (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Fibroblast growth factor 2 (fgf2), by STEMCELL (1 mentions)
Horse serum, by Merck Group (1 mentions)
H1138, by Merck Group (1 mentions)
Epidermal growth factor (egf), by STEMCELL (1 mentions)
D5796, by Merck Group (1 mentions)
Neurobasal, by Thermo Fisher Scientific (1 mentions)
100 18b 1mg, by Thermo Fisher Scientific (1 mentions)
Nucleofector 2b device, by Lonza (1 mentions)
5 protocols using scc008
1
Culturing Human Cell Lines for Research
2
Glioma and Neural Progenitor Cell Cultures
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TS603, TS667, TS543 were obtained from Memorial Sloan Kettering Cancer Center (MSKCC). SU-AO3, NCH612 glioma lines were described previously [23 (link)]. The patient-derived IDH1-mutant glioma lines were routinely subjected to panel sequencing analysis [26 (link)] and immunoblotting to confirm the expression of mutant IDH1 (R132H). Glioma lines were maintained as neurospheres in NeuroCult (StemCell Technologies) with 2 µg/ml heparin sulfate (Sigma), 20 ng/ml of EGF (StemCell Technologies), and FGF2 (StemCell Technologies). PC12 cells (provided by Dr. Agarwal, Heidelberg University; Dr. Chang, Gachon University) were maintained in DMEM (D5796, Sigma) supplemented with 10% horse serum (H1138, Sigma) and 1% penicillin–streptomycin. Human neural progenitor cells (SCC008, Sigma) were maintained in DMEM (D5796, Sigma) supplemented with 10% fetal bovine serum (Gibco) and 1% penicillin–streptomycin. All cells were grown in a 5% CO2 humidified incubator at 37 °C.
3
Human Neural Progenitor Cell Culture
4
Culturing and Transfecting Human Neural Progenitor Cells
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ReNcell VM neural progenitor cells (Merck Millipore SCC008) were used in this study as an in vitro model of hNSC. They were grown either as neurospheres or as adherent monolayer on laminin coated plasticware in the following culture medium: 50% DMEM:F12 (GIBCO 11554546), 50% Neurobasal (GIBCO 21103049), 0.5% Penicillin-Streptomycin (Sigma-Aldrich P4333-100ML), 1% N2 supplement (GIBCO 17502048), 2% B27 NeuroMix (GIBCO 17504044), heparin (5μg/ml, Sigma-Aldrich 9041-08-1). Human Epidermal Growth Factor (hEGF, GIBCO AF-100-15-1MG) and human Fibroblast Growth Factor (hFGF, GIBCO 100-18B-1MG) were added to the cell culture medium just before use, both at a concentration of 20 ng/ml. Growth medium was replaced with fresh medium every 2 days.
Transfection of hNSC was performed by electroporation using a NucleofectorTM 2b Device (Lonza) with NucleofectorTM Kits for Mouse Neural Stem Cells as per manufacturer instructions (Lonza VPG-1004).
Transfection of hNSC was performed by electroporation using a NucleofectorTM 2b Device (Lonza) with NucleofectorTM Kits for Mouse Neural Stem Cells as per manufacturer instructions (Lonza VPG-1004).
5
Gymnotic Uptake of ASO in ReNcells VM
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Prior to proceeding with the gymnotic uptake, ASO was dissolved in nuclease-free H2O and the concentration of ASO solution was determined by OD260 nm absorbance. ReNcells VM (EMD Millipore, SCC008) were grown in laminin coated flasks in complete NSC medium. Complete NSC medium was prepared by supplementing maintenance medium (Millipore, SCM005) with bFGF (EMD Millipore, GF003) and EGF (EMD Millipore, GF144), to a final concentration 20 ng/mL each. Cells were detached from the flask by incubation with accutase (Millipore, SCR005), resuspended in prewarmed maintenance medium and pelleted at 300 × g by centrifugation. The supernatant was removed by aspiration and the cells was resuspended in complete NSC medium to ~1 × 106/mL. In all, 0.5 mL of cell suspension (~5 × 105 cells) was added to each well of an Ultra-Low Attachment Costar 24 well plate (Corning, 3473) containing ASO. The cells were mixed gently with ASO and were incubated at 37 °C, 5% CO2 for 72 h before harvesting.
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