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I have been told that setting more than one discriminator on the Elite (or
perhaps any machine) can lead you into trouble, because the instrument uses
logic when dealing with multiple discriminators that can wind up giving
counter-intuitive effects.
I have found that setting a discriminator on a fluorescent signal,
particularly a red signal that may be present in particles unrelated to
those carrying other labels, can lead to odd results. Remember that data
collection on all parameters is triggered only when a signal rises above
the discriminator on the parameter you choose. You need to play around with
red channel gains and discriminator level to make sure that the data
rejected by the discriminator is not the data that carries your FITC signal
(as is obviously happening at the moment).
Unless there is a very good reason for doing otherwise, I have found it
safer overall to discriminate only on a light scatter parameter, every
particle scatters some light at least and junk is usually the small stuff.
Dean R. Hewish, Cell Biologist & Flow Cytometrist.
CSIRO Biomolecular Engineering, 343 Royal Parade, Parkville, 3052,
Victoria Australia.
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