The flow of information was more of the FACSort than the Turbo variety with respect to rate, and now that the trickle has stopped, I guess it is time to share what I got. All responses were informative, some quite entertaining. The general consensus is that viability, not unlike life itself, is pressure sensitive: some can take it better than others. All agree that viability of murine lymphoid and bone marrow cells is excellent when sorted at 30 to 33 psi with the BD TurboSort. Functionality, and colony forming potential of progenitors is uncompromised. One brave soul found that at 45 psi, using a 70 micron tip, only 6% of the lymphoid population and 8% of the granulocyte population were PI positive after Turbo sorting. With respect to human myeloma cell lines there was no reported effect on viability, and only a small difference in colony forming units in number and size. Turbo sorted(human) CD34 positive cells had the same viability and colony forming potential as non-Turbo sorted cells. There were a few responses from the MoFlo camp: viability at 60 psi is excellent, with only 2% of the population staining PI positive after sorting, and 7% staining PI positive at 80 psi (mouse bone marrow). One respondent indicated that using the MoFlo, single bacteria cloned out at 100%, and bull semen gave pregnancy rates similar to unsorted material. Ok .... The exceptions are fragile cells: ie destined for apoptosis heaven [ should use less than 22 psi ], or stimulated (spenocytes) [22 psi ], dendritic and microglial cells [12 psi]. These cell types benefit from lower sort pressures and larger nozzle tips. Selection of nozzle tip diameter is important if not critical for good results: large, more fragile cells show less mortality with larger diameter tips. Nozzle tip design ... I have to assume the MoFlo uses a different design from BD ... also influences viability at increased pressures (kinda intuitive, but the implication is that nozzle tip selection has more effect on viability than pressure for most cell types). There is the question of TurboSort use with the autoclone unit: Why do this at high speed since especially in 96 well plates, far more cells will go to the waste as the unit travels between wells. There may be some gain in sorting for very rare events or sorting large numbers of cells into 24 well plates however. And finally, last but certainly not least, many agree that the biggest factor in cell viability ... in most cases ... is cell prep, not sorter pressure. Recommendations include the use of HEPES buffered saline over phosphate buffered saline since the former is less susceptible to changes pH with exposure to the atmosphere. Verifying the osmolarity of the sheath fluid is also recommended. And that about sums it up. PS: don't ask me, I don't have a high speed sorter ... maybe some day I will, judging from the responses in this survey. Again, sincere thanks to all who responded, Claude
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