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I think your idea of getting a volume measurement is very interesting.
However, I believe in reality it would be quite difficult to make it very
accurate. This is for two or three reasons. First, time of flight measures
not only the particle size but the laser beam configuration. You have to do
a deconvolution of the laser beam to get an accurate measure of size. For
smallish cells, this will be prone to inaccuracy. A tightly focused laser
beam (a few microns) makes this easier. Second, since volume is the cube of
diameter, a rather small change in diameter is reflected in a large change in
volume. Conversely, a small change in volume reflects a very small change in
diameter, so small that I suspect it would be difficult to measure accurately
and reproducibly using time-of-flight. Thirdly, if cells are not precisely
spherical (e.g. elongated), the time-of-flight measurement would be skewed.
We tried to use time-of-flight on the old Coulter MDADS system and found it
not to be very reliable for our cells because of these reasons. Many of
these problems disappear or become less important, when using large cells,
such as plant protoplasts.
Maybe you or David Galbraith can prove me wrong and make this workable.
Certainly it would be nice to have something better than forward angle light
scatter!
Mike Fox
Colorado State University
Fort Collins, CO
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