NCH421K Cells

Categories: Brain cancer cell lines Description: This cell line was established by Christel Herold-Mende from glioblastoma tissue. Organism: Human Tissue: Brain Disease: Glioblastoma Synonyms: NCH421k Age: 66 years Gender: Male Ethnicity: Caucasian Growth Properties: Spheroid culture Citation: NCH421K (Cytion catalog number 300118) Biosafety Level: 1 Ncbi_ Taxid: 9606.0 Cellosaurus Accession: CVCL_x910 Tumorigenic: Yes Culture Medium: DMEM:Ham's F12 (1:1), w: 3.1 g/L Glucose, w: 2.5 mM L-Glutamine, w: 15 mM HEPES, w: 0.5 mM Sodium pyruvate, w: 1.2 g/L NaHCO3 (Cytion article number 820400a) Supplements: Supplement the medium with 10% FBS, 5 mg/L Heparin, 20 ng/mL bFGF, 20 microgram/L EGF, 5 mg/L Insulin, 100 mg/L Transferrin, 5,2 microgram/L Na-selenit, 6,3 microgram/L Progesteron, 161,1 microgram/L Putrescin, 50 mg/L Hydrocortinson Doubling Time: 35 to 40 hours Subculturing: For subculturing spheroid cultures, begin by mechanically dissociating the spheroids through pipetting up and down 5 to 10 times using an Eppendorf pipette with 1000 µl filter tips. After this, centrifuge the mixture at 300g for 5 minutes at room temperature to pellet the cells. Discard the supernatant and resuspend the cell pellet in fresh culture medium. Finally, transfer the resuspended cells into new culture vessels to promote further spheroid formation. This approach ensures efficient spheroid breakdown and readies them for continued growth in a new environment. Seeding Density: 1 to 2 x 105 cells/ml Fluid Renewal: 2 to 3 times per week Post Thaw Recovery: Please allow the cells to recover from the freezing process for at least 24 to 48 hours. Freeze Medium: As a cryopreservation medium, use 50% basal medium + 40% FBS + 10% DMSO, or CM-1 (Cytion catalog number 800100), which includes optimized osmoprotectants and metabolic stabilizers to enhance recovery and reduce cryo-induced stress. Thawing And Culturing Cells: Confirm that the vial remains deeply frozen upon delivery, as cells are shipped on dry ice to maintain optimal temperatures during transit. Upon receipt, either store the cryovial immediately at temperatures below -150°C to ensure the preservation of cellular integrity, or proceed to step 3 if immediate culturing is required. For immediate culturing, swiftly thaw the vial by immersing it in a 37°C water bath with clean water and an antimicrobial agent, agitating gently for 40-60 seconds until a small ice clump remains. Perform all subsequent steps under sterile conditions in a flow hood, disinfecting the cryovial with 70% ethanol before opening. Carefully open the disinfected vial and transfer the cell suspension into a 15 ml centrifuge tube containing 8 ml of room-temperature culture medium, mixing gently. Centrifuge the mixture at 300 x g for 3 minutes to separate the cells and carefully discard the supernatant containing residual freezing medium. Gently resuspend the cell pellet in 10 ml of fresh culture medium. For adherent cells, divide the suspension between two T25 culture flasks, for suspension cultures, transfer all the medium into one T25 flask to promote effective cell interaction and growth. Adhere to established subculture protocols for continued growth and maintenance of the cell line, ensuring reliable experimental outcomes. Sterility: Mycoplasma contamination is excluded using both PCR-based assays and luminescence-based mycoplasma detection methods. To ensure there is no bacterial, fungal, or yeast contamination, cell cultures are subjected to daily visual inspections. Safety Precautions: When planning to store a cryovial in liquid nitrogen for future thawing, it is mandatory to adhere to stringent safety measures. Appropriate protective gloves and clothing are essential, and the use of a face mask or safety goggles is required during the transfer of frozen samples to or from the liquid nitrogen tank. This is to mitigate the risk of injury from potential cryovial explosions upon removal, which can result in the projection of sharp fragments. Warranty: We stand by the promise of delivering products with high cell viability and robust culture performance. To achieve the best results, please make sure you follow the storage and culture instructions detailed in the product information sheet closely. Your adherence to these guidelines is key to success. Subject To Third- Party Agreements: Please note that this cell line is not available under a standard Cytion MTA, as it requires a third-party agreement and/or is subject to negotiation with the original licensor. Required Product 1: 820400a Required Product 3: 860015.0 Required Product 4: 830100.0