Preserving high quality RNA for post-cell-sort sequencing in fixed cells can be achieved using a zinc-buffer fixation protocol.

Information posted March 27, 2019 on the Purdue-administered flow cytometry bulletin board by Dr. Roxana del Rio-Guerra says -

"We, the (ABRF-) Flow Cytometry Research Group just presented our work at the last ABRF meeting (poster 129) about the different fixative methods (including commercially available products for that purpose) and the purity, quality and yield of RNA after sorting cells. We tested: paraformaldehyde (2 and 4%), formaldehyde (1 and 4 %), alcohols (ethanol and methanol), Zinc buffer, as well as commercially available kits such as CytoFix/Cyto perm and FACSLyse (from BD), and IC fix/perm buffer (eBio/ThermoFis). Each of these protocols was performed by two different Flow core labs at two different institutions and all samples sent to a general lab for evaluation of the integrity/quantity of RNA. Among all those protocols, the Zinc Buffer protocol (Christensen, R, et al. Curr Protoc Cytom 2011, Chapter 7: Unit 7.40. PMID: 21732310) gave us a very promising results with near to 8.0 on the RNA integrity number. For our work, we first stained cells only with a viability dye and then fix-sort-fix."

In a follow-up message Dr. del Rio-Guerra adds -

"Also, on ref from Curr Prot Cyt, we changed from Zinc Acetate to 17.16 mM zinc-trifluoroacetate since this was shown to better preserve the quality of RNA (ref PMID: 17576663) We tested: after fix-perm (with the diff fixatives mentioned), sorted cells were collected on RLT plus buffer (Qiagen 1053393). With the exception of Zn buffer, quality of RNA was very poor. The results presented at the last ABRF meeting are preliminary, but the Zn buffer protocol was the one reproducible in two different core labs and with the best quality of RNA. We will continue working on this project."

If you have access to Current Protocols in Cytometry , you can access the zinc-buffer fixation protocol directly. The Flow Cytometry Facility can provide the protocol in PDF by request.