人大脑皮层神经元Cell Specification
The tissue of the central nervous system is made up of two classes of cells that may be broadly
categorized as neurons and glia. Neurons are anatomic, functional, and trophic units of the brain
[1]. Despite great variability in size and shape, all neurons share common morphological
features, the key elements of a highly complex communication network. Neurons are
dynamically polarized cells that serve as the major signaling unit of the nervous system. The
human brain contains about 1 x 1011 neurons and each are able to contact at least 10,000 other
neurons [2].
HN-mb from ScienCell Research Laboratories are isolated from human midbrain
(mesencephalon). HN-mb are cryopreserved at P0 and delivered frozen. Each vial contains >1 x
106
cells in 1 ml volume. HN-mb are characterized by immunofluorescence with antibodies
人大脑皮层神经元specific to neurofilament, MAP2, and β-tubulin III. HN-mb are negative for HIV-1, HBV, HCV,
mycoplasma, bacteria, yeast, and fungi. HN-mb are guaranteed to further culture under the
conditions provided by ScienCell Research Laboratories; however, HN mb are not recommended
for expanding or lon term cultures since the cells do not proliferate in culture.
Recommended Medium
It is recommended to use neuronal medium (NM, Cat. #1521) for culturing HN-mb in vitro.
Product Use
HN-mb are for research use only. They are not approved for human or animal use, or for
application in in vitro diagnostic procedures.
Storage
Upon receiving, directly and immediately transfer the cells from dry ice to liquid nitrogen and
keep the cells in liquid nitrogen until they are needed for experiments.
Shipping
Dry ice.
References
[1] Parent A. (1996) “Neurons.” In Carpenters Human Neuroanatomy (9th ed., pp131-198). Quebec: Williams &
Wilkins.
[2] Alberts B, Bray D, Lewis J, Raff M, Roberts M, Watson JD. (1989) Molecular Biology of the Cell (2nd ed.).
New York: Garland.
Instructions for culturing cells
Caution: Cryopreserved cells are very delicate. Thaw the vial in a 37oC water bath
and return the cells to culture as quickly as possible with minimal handling!
Initiating the culture:
1. Prepare a poly-L-lysine-coated culture vessel (2 μg/cm2
, T-25 flask is recommended).
Add 5 ml of sterile water to a T-25 flask and then add 5 μl of poly-L-lysine stock solution
人大脑皮层神经元(10 mg/ml, Cat. #0413). Leave the vessel in a 37oC incubator overnight (or for a
minimum of one hour). Rinse the poly-L-lysine-coated vessel twice with sterile water
prior to use.
Note: It is important that these cells are plated in poly lysine coated culture vessels to
promote cell attachment.
2. Prepare complete medium. Decontaminate the external surfaces of medium bottle and
medium supplement tubes with 70% ethanol and transfer them to a sterile field.
Aseptically transfer supplement to the basal medium with a pipette. Rinse the supplement
tube with medium to recover the entire volume.
3. Add complete medium to the culture vessel. Leave the vessel in the sterile field and
proceed to thaw the cryopreserved cells.
4. Place the frozen vial in a 37oC water bath. Hold and rotate the vial gently until the
contents completely thaw. Promptly remove the vial from the water bath, wipe it down
with 70% ethanol, and transfer it to the sterile field. Carefully remove the cap without
touching the interior threads.
5. Gently resuspend and dispense the contents of the vial into the poly-L-lysine-coated
culture vessel. A seeding density of 10,000-50,000 cells/cm2 is recommended, with an
optimal range of 20,000-25,000 cells/cm2
.
Note: Dilution and centrifugation of cells after thawing are not recommended since these
actions are more harmful to the cells than the effect of residual DMSO in the culture.
6. Replace the cap or lid of the culture vessel and gently rock the vessel to distribute the
cells evenly. Loosen cap, if necessary, to allow gas exchange.
7. Return the culture vessel to the incubator.
8. For best results, do not disturb the culture for at least 16 hours after the culture has been
initiated. Refresh culture medium the next day to remove residual DMSO and unattached
cells, then every other day thereafter.
Maintaining the culture:
1. Refresh supplemented culture medium the next morning after establishing a culture from
cryopreserved cells.
2. Change the medium every two to three days thereafter.
It is not recommended that neurons be subcultured beyond their initial plating.
人大脑皮层神经元Caution: Handling human derived products is potentially biohazardous. Although each cell
strain tests negative for HIV, HBV, and HCV DNA, diagnostic tests are not necessarily 100%
accurate, therefore, proper precautions must be taken to avoid inadvertent exposure. Always
wear gloves and safety glasses when working with these materials. Never mouth pipette. We
recommend following the universal procedures for handling products of human origin as the
minimum precaution against contamination [1].
[1] Grizzle WE, Polt S. (1988) “Guidelines to avoid personal contamination by infective agents in research
laboratories that use human tissues.” J Tissue Cult Methods. 11: 191-9.