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== FACS Sort == :*DAPI stained or DAPI/NeuN stained (Note: Rnase Inhibitor was added during staining) human nuclei from Gwen (J. Chun Lab): :**Added ~1ul DAPI just prior to FACS sort on the Aria (Scripps) :**Nuclei were maintained on ice :**Added 50ul of sort buffer containing 0.5% FAF-BSA to low-bind 1.5ml eppendorf tube for collection :**Gated nuclei on DAPI positive and shifted gate slightly along the FSC axis to exclude the smallest nuclei :**Sorted 150,000 DAPI+ nuclei and 75,000 NeuN+ nuclei into separate collection tubes (final volume of ~1000ul) :**Note that the DAPI positive showed banding pattern. For NeuN stained we selected only the lowest band (single nuclei) and avoided the upper bands that could be multiples. This was not done for the DAPI+ alone :**Maintained on ice. Centrifuged 2000rpm at 4C for 10 minutes. Removed supernatant down to ~10ul. Resuspended pellet with ~20ul 0.5%BSA/PBS using p1000 by pipetting about 10 times. :**Counted nuclei using BioRad automatic cell counter: '''1200 nuclei/ul for DAPI+ and 1100 nuclei/ul for NeuN+''' :**Adjusted [nuclei] to 750 nuclei/ul for both (final volume of 45-50ul)
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