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Product Name Human Neuron CellsCatalog Number HNC001Size 1mLStorage Liquid NitrogenCell Number: Frozen Vial (500.000 per vial)Product Format: Frozen
Caution: Proper precautions must be taken to avoid exposure. Always wear proper protective equipment(Gloves, safety glasses, etc.) when handling these materials. We recommend following the universalprocedures for handling products of human origin as the minimum precaution against contamination.The listed dilutions are for recommendation only and the ender users should optimize the final conditions.General InformationHNCs (HNC001) are initiated by digestion of minced brain cortical tissue with collagenase. HNCsare separated from the mixture of cell populations and offered at passage 3 in a frozen vial. HNCs GrowthMedium (contains 10% serum and growth supplements, HNM 001) is recommended for cell culture andthese cells have a minimum average population doubling capacity > 8 when cultured following thedetailed protocol described below).Characterization of the cellsThe cells stain positively for a number of neuronal markers:Neurofilament protein PositiveNeuron specific enolase (NSE) PositiveThe cells are negative forGlial fibrillary acidic protein (GFAP) NegativeMyelin basis protein (MBP) NegativeHNCs are negative for HIV-1, HBV, HCV, and mycoplasma.Product Use: HNCs are for research use only.Shipping: Frozen Vials in Dry Ice packagesHandling of Arriving CellsWhen you receive the cells in a frozen vial, you can transfer the vial of cells into a -80ºC freezer for shortperiod storage or a liquid nitrogen tank for long term storage. Thaw the cells in a 37°C water bath, andthen transfer the cells into a T25 flask pre-coated with Alphabiocoat solution as described in details inSubcultureProtocol
Subculture ProtocolA) Pre-coating of T25 flasks: Add 2ml of Alphabiocoat Solution into one T25 flask and make sure wholesurface of the flask is covered with the coating solution. Five minutes later, dispose excessiveAlphabiocoat Solution by aspiration and the flask is ready to be used (no need for overnight incubationwhen using Alphabiocoat Solution).B) Rinse the cells in T25 flask with 5ml HBSS (Room Temperature, RT) twice.C) Add 2ml of Trypsin/EDTA (RT) (cAP-23) into one T25 flask (make sure the whole surface of the T25flask is covered with Trypsin/EDTA), and gently dispose the excessive Trypsin/EDTA solution within60 seconds with aspiration.D) Leave the T25 flask with the cells at 37C for extra 1-2 minute (the cells usually will detach from thesurface within 1-2 minutes). You can monitor the cells under microscope and when most of cells becomerounded up, hit the flask against the bench surface, and the cells will move on the surface of the flaskwhen monitoring under microscope.E) Add 5ml Trypsin Neutralization Buffer and spin the cells down with 800g for 5 minutes.F) Re-suspend the cell pellet with 10 – 15ml of HNCs Growth Medium and the cell suspension istransferred directly into 2 or 3 pre-coated T25 flasks (5ml each, and the cells are sub-cultured at 1 : 2 to1 : 3 ratios)G) Change medium every 2-3 days and cells usually become confluent within 7 days (when split at a 1:4ratio).