We are trying to isolate RNA from astrocyte after FACS. We use 7AAD to exclude the dead cell which was without clearly boundary of positive and negtive. we tried to load the cells either in the facs buffer or directly into RLY lysis buffer or trizol reagents. The cell numbers was 10^4-10^5, the concentration of RNA was always around 20 pg/ul(total 240pg). Should we change to use zoombie or FVS to increase the viability of the cells? any other advice ?