Although I do as little sorting as possible, I have been following this thread with a great deal of interest. It is inspiring to see people abandoning the convenient "my sorter can sort faster than your sorter" mantra and asking which of the differing physical parameters of "high speed" and "low speed" sorting regimes may be responsible for loss of cell viability or function. Pressure is certainly a candidate. It seems intuitively unlikely that exposure to loud noise at 60 or 100 kHz should be harder on cells than exposure to loud noise at 10 to 40 kHz. And I don't think that traveling 30 m/sec is worse than traveling 10 m/sec; the g-force coming out the nozzle shouldn't be that high (although I haven't done the calculation - it's been a long time since physics class). People used to talk about shear stress affecting cells, but with sheath flow, the cells shouldn't be subjected to big-time shear stress. So it's probably pressure, or changes in pressure. I remember that when the first of the Livermore high-speed sorters went on the air, their users were concerned primarily with sorting chromosomes, which were unaffected by the high speed and pressure, and not cells, which didn't survive a trip through the sorter. There was talk of cells "blowing up", experiencing a severe case of decompression sickness or "the bends" at the cellular level. That is still probably a good model for cell damage; maybe the sublethal effects reflect "rapture of the drops". Physiologists have learned a lot about how to keep the large cell aggregates which comprise divers and astronauts alive after exposure to extremes of pressure; it's usually rapid decompression which causes problems, and the solution typically involves slowing down decompression and/or modifying the ambient gas mixture to minimize the formation of microbubbles. It might be a good idea to look at cells subjected to high pressure in bulk, and compare the effects of rapid and slow decompression on viability and function. If cells decompressed slowly survive and function better than those decompressed rapidly, it would be advisable to look at different gas mixtures, since it is probably impractical to slow decompression as cells go through the nozzle into air at high pressure - and speed. Dredging up memories again, I remember Linus Pauling extolling the virtues of xenon as an anesthetic - perhaps it would make a good sheath drive gas. That would certainly force changes in the rate structure of sorting facilities. But, as I said, I sort as little as possible, so I'm not under pressure... -Howard
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