Transfer a mouse brain enriched immune pellet from a 15-milliliter tube to a round bottom 96-well plate. After centrifuging the plate at 500 G for five minutes, Remove the supernatant by flicking. Resuspend the cells in 100 microliters of FACS buffer, then transfer 10 microliters from each well into the control wells.
Centrifuge at 500 G for five minutes. After removing the supernatant from the centrifuged cells, add 50 microliters of double concentration FC block. Incubate for 10 minutes on ice.
Now add 50 microliters of the antibody master mix at double the final desired concentration and incubate it on ice in the dark. Then, add 200 microliters of the FACS buffer to each well and centrifuge at 500 G for five minutes. Discard the supernatant by flicking, then wash the cells in 300 microliters of FACS buffer.
Centrifuge and resuspend the cells in 200 microliters of FACS buffer. Finally, transfer the suspension to a flow sort tube containing 300 microliters of FACS buffer.
