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Apologies are unnecessary, now that Mario and so many people are running
three beams. When we didn't have dyes with a variety of Stokes shifts, like
the phycobiliproteins and tandems, multiple excitation beams provided the
only practical approach to multicolor immunofluorescence measurement. We
can now use fewer beams, but, as the number of colors per beam increases,
the problem of compensation becomes harder to solve with either hardware or
software. It's hard to build systems deriving 4 or more separated beams
when you get the beams from big lasers, but is considerably easier when you
use diode or solid state lasers, now eminently suitable for green, red, and
infrared excitation, and probably available for UV through blue within a few
years. If our appetites for more and more immunofluorescence colors keep
increasing, we'll have to consider both approaches.
Of course, the original Block system (1974) derived five separated beams
from one arc lamp, but, then again, we weren't doing immunofluorescence.
-Howard
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CD-ROM Vol 3 was produced by Monica M. Shively and other staff at the
Purdue University Cytometry Laboratories and distributed free of charge
as an educational service to the cytometry community.
If you have any comments please direct them to
Dr. J. Paul Robinson, Professor & Director,
PUCL, Purdue University, West Lafayette, IN 47907.
Phone: (765)-494-0757;
FAX(765) 494-0517;
Web