大鼠表皮角质细胞-新生-细胞株/菌种-试剂-生物在线
大鼠表皮角质细胞-新生

大鼠表皮角质细胞-新生

商家询价

产品名称: 大鼠表皮角质细胞-新生

英文名称: Rat Epidermal Keratinocytes-neonate

产品编号: XY2100

产品价格: 0

产品产地: 中国/美国

品牌商标: XYbscience

更新时间: 2023-08-17T09:55:27

使用范围: null

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大鼠表皮角质细胞-新生Cell Specification
The epithelial layer of the skin provides an essential function as a protective barrier against insult
from the outside environment. The major cell type in this layer is keratinocytes which consists
around 85% of living epidermal cells. Keratinocytes are the cells of stratified squamous
epithelia. It is so named because its most abundant protein is keratin. Keratinocyte divide and are
thus generated in the basal layer of the epidermis; this division is followed by a programmed
death as they produce keratin and move up to the surface of the epidermis. Keratinocyte
proliferation, differentiation, and apoptosis are complex and carefully choreographed processes
[1]. Besides the protective function, keratinocytes also play a central role in tissue homeostasis,
wound healing, cancers, and skin-based gene-therapy. Keratinocytes express adhesion molecules
and cytokines indicating their participation in skin innate immunity and homeostasis in vivo [2,
3].
REK-n from ScienCell Research Laboratories are isolated from neonate rat epidermal tissue.
大鼠表皮角质细胞-新生REK-n are cryopreserved on primary culture and delivered frozen. Each vial contains >5 x 105
cells in 1 ml volume. REK-n are characterized by immunofluorescent method with antibodies to
cytokeratine-18 and -19. REK-n are negative for mycoplasma, bacteria, yeast and fungi. REK-n
are guaranteed to further expand for 15 population doublings in the condition provided by
ScienCell Research Laboratories.
Recommended Medium
It is recommended to use Keratinocyte Medium (KM, Cat. No. 2101) for the culturing of REK-n
in vitro.
Product Use
REK-n are for research use only. It is not approved for human or animal use, or for application in
in vitro diagnostic procedures.
Storage
Directly and immediately transfer cells from dry ice to liquid nitrogen upon receiving and keep
the cells in liquid nitrogen until cell culture needed for experiments.
Shipping
Dry ice.
Reference
[1] Eckert, R. L., Efimova, T., Dashti, S. R., Balasubramanian, S., Deucher, A., Crish, J. F., Sturniolo, M. and Bone,
F. (2002) Keratinocyte survival, differentiation, and death: many roads lead to mitogen-activated protein kinase.
J Investig Dermatol Symp Proc 7(1):36-40.
[2] Song, P. I., Park, Y. M., Abraham, T., Harten, B., Zivony, A., Neparidze, N., Armstrong, C. A. and Ansel, J. C.
(2002) Human keratinocytes express functional CD14 and toll-like receptor 4. J Invest Dermatol 119(2):424-32.
[3] de Panfilis, G., Semenza, D., Lavazza, A., Mulder, A. A., Mommaas, A. M. and Pasolini, G. (2002)
Keratinocytes constitutively express the CD95 ligand molecule on the plasma membrane: an in situ
immunoelectron microscopy study on ultracryosections of normal human skin.
Br J Dermatol. 147(1):7-12.
ScienCell
Research Laboratories
TM
大鼠表皮角质细胞-新生Instruction for culturing cells
Caution: Cryopreserved cells are very delicate. Thaw the vial in a 37oC waterbath
and return them to culture as quickly as possible with minimal handling!
Set up culture after receiving the order:
1. Prepare a poly-L-lysine coated flask (2 μg/cm2
, T-75 flask is recommended). Add 10 ml
of sterile water to a T-75 flask and then add 15 μl of poly-L-lysine stock solution (10
mg/ml, ScienCell cat. no. 0413). Leave the flask in incubator overnight (minimum one
hour at 37oC incubator).
2. Prepare complete medium: decontaminate the external surfaces of medium and medium
supplements with 70% ethanol and transfer them to sterile field. Aseptically open each
supplement tube and add them to the basal medium with a pipette. Rinse each tube with
medium to recover the entire volume.
3. Rinse the poly-L-lysine coated flask with sterile water twice and add 20 ml of complete
medium to the flask. Leave the flask in the hood and go to thaw the cells.
4. Place the vial in a 37oC waterbath, hold and rotate the vial gently until the contents are
completely thawed. Remove the vial from the waterbath immediately, wipe it dry, rinse
the vial with 70% ethanol and transfer it to a sterile field. Remove the cap, being careful
not to touch the interior threads with fingers. Using a 1 ml eppendorf pipette gently resuspend
the contents of the vial.
5. Dispense the contents of the vial into the equilibrated, poly-L-lysine coated culture
vessels. A seeding density of 5,000 cells/cm2
is recommended.
Note: Dilution and centrifugation of cells after thawing are not recommended since these
actions are more harmful to the cells than the effect of DMSO residue in the culture. It is
also important that cells are plated in poly-L-lysine coated flask that promotes cell
attachment and growth.
6. Replace the cap or cover, and gently rock the vessel to distribute the cells evenly. Loosen
cap if necessary to permit gas exchange.
7. Return the culture vessels to the incubator.
8. For best result, do not disturb the culture for at least 16 hours after the culture has been
initiated. Change the growth medium the next day to remove the residual DMSO and
unattached cells, then every other day thereafter.
Maintenance of Culture:
1. Change the medium to fresh supplemented medium the next morning after establishing a
culture from cryopreserved cells.
2. Change the medium every three days thereafter, until the culture is approximately 70%
confluent.
3. Once the culture reaches 70% confluence, change medium every other day until the
culture is approximately 90% confluent.
Subculture:
1. Subculture the cells when they are over 90% confluent.
2. Prepare poly-L-lysine coated cell culture flasks (2 μg/cm2
).
3. Warm medium, trypsin/EDTA solution (T/E, cat. no. 0103), trypsin neutralization
solution (TNS, cat. no. 0113), and DPBS (Ca++ and Mg++ free, cat. no. 0303) to room
temperature. We do not recommend warming the reagents and medium at 37oC
waterbath prior to use.
4. Rinse the cells with DPBS.
5. Add 5 ml of DPBS first and then 5 ml of trypsin/EDTA solution into flask (in the case of
T-75 flask); gently rock the flask to make sure cells are covered by trypsin/EDTA
solution; incubate the flask at 37oC incubator for 3 to 5 minutes or until cells are
completely rounded up (monitored with inverted microscope). During incubation, prepare
a 50 ml conical centrifuge tube with 5 ml of fetal bovine serum (FBS, cat. no. 0500);
transfer trypsin/EDTA solution from the flask to the 50 ml centrifuge tube (a few percent
of cells may detached); continue incubate the flask at 37oC for 1 or 2 minutes more (no
solution in the flask at this moment); at the end of trypsinization, one hand hold one side
of flask and the other hand gently tap the other side of the flask to detach cells from
attachment; check the flask under inverted microscope to make sure all cells are
detached, add 5 ml of trypsin neutralization solution to the flask and transfer detached
cells to the 50 ml centrifuge tube; add another 5 ml of TNS to harvest the residue cells
and transfer it to the 50 ml centrifuge tube. Examine the flask under inverted microscope
to make sure the cell harvesting is successful by looking at the number of cells left
behind. There should be less than 5%.
Note: Use ScienCell Research Laboratories’ trypsin/EDTA solution that is optimized to
minimize the killing of the cells by over trypsinization.
6. Centrifuge the 50 ml centrifuge tube (harvested cell suspension) at 1000 rpm (Beckman
Coulter Allegra 6R centrifuge or similar) for 5 min; re-suspend cells in growth medium.
大鼠表皮角质细胞-新生7. Count cells and plate cells in a new, poly-L-lysine coated flask with cell density as
recommended.
Caution: Handling animal derived products is potentially biohazardous. Always wear gloves and safety glasses
when working 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, W. E., and Polt, S. S. (1988) Guidelines to avoid personal contamination by infective agents in research
laboratories that use human tissues. J Tissue Culture Methods. 11(4).