hTert Immortalized GFP-Human Brain Microglia Cells

Frozen Vials in a Dry Ice Package.

Frozen vial

General Information# Human brain microglia cells (HBMgs, cAP-0040) are isolated from healthy human brain tissue. hTert-Immortalised GFP-Human Brain Microglia Cells (HBMgs-hTert-GFP) are selected from HBMgs infected with lentiviruses expressing hTert and GFP with puromycin. HBMgs can be cultured in long-term (tested >20 passages). It is recommended to use Microglia Cell Medium (MCM, cAP-37; ) for the culturing of HBMgs.
Characterization of the Cells#
1.: Cytoplasmic F4/80 Positive
2.:CD68 Positive
3.:HBMgs are negative for HIV-1, HBV, HCV, and mycoplasma.
Product Usage# The cells are offered for Research Use Only.
Handling of Arriving Cells# When you receive the cells in a frozen vial, you can transfer the vial of cells into a -80C freezer for short period storage or a liquid nitrogen tank for long-term storage. Thaw the cells in a 37C water bath, and then transfer the cells into a T25 flask pre-coated with poly-L-lysine as described in following details.
Protocols for Thawing and maintaining the cells#
A):Prepare a poly-L-lysine coated flask (2ug/cm2 , T-25 flask is recommended). Add 5 ml of sterile water to a T-25 flask and then add 9ul of poly-L-lysine stock solution (cAP-38, 10mg/ml). Leave the flask in an incubator overnight (minimum one hour ata 37C incubator).
B):Rinse the poly-L-lysine coated flask with sterile water twice and add 7 ml of complete medium to the flask. Leave the flask in the hood and go to thaw the cells.
C):Warm MCM (cAP-37) before thawing the cells.
D):Place the vial in a 37C water bath, hold and rotate the vial gently until the contents are completely thawed. Remove the vial from the water bath 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 1 ml Eppendorf pipette gently resuspend the contents of the vial.
E):Dispense the contents of the vial into the equilibrated, poly-L-lysine coated culture vessels. A seeding density of more than 10, 000 cells/cm2 is recommended. Note: Since these cells adhere very strongly to tissue culture dishes, trypsinization after attachment is incomplete. Therefore, it is recommended to plate the cells in the experimental format called for and use them directly for the experiment, instead of trying to subculture them. The cells can also be maintained on ultra low binding dishes (Corning) or on Poly-L-lysin plates.
F):Replace the cap or cover, and gently rock the vessel to distribute the cells evenly. Loosen cap if necessary to permit gas exchange.
G):Return the culture vessels to the incubator (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 by centrifuging the unattached cells down, resuspending the cells with 5ml fresh medium, and add the cells back to the flask. Then change the medium every other day thereafter).
H):Change the medium every two to three days thereafter.