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Our standard sample tube for dye laser setup includes APC beads (Flow
Cytometry Standards Corp.), Texas beads (FCSC) and FITC beads (B_D
calibrites). The Texas and APC beads (i) have fluorescence in the range
we normally see with immunofluorescent staining and (ii) they can be
used to set the Texas/APC compensation if we're setting up for both
fluorochromes. The FITC beads do not excite with the dye laser so they
give a good negative background level. It is important to have
negatives in your sample because there are ways to get droplet drive
ripple or other extraneous signals into the Texas or APC detectors. If
you are starting with the alignment way out, it's handy to know if the
signals you are seeing are related to fluorescence. The FITC beads can
also be used at the same time to double check that the laser1 alignment
is still ok.
You should also check your obscuration bar while looking at both laser1
and laser2 signals; (presumably) your lasers have separated stream
intercepts so the bar must cover reflections from both.
\ / < Flow Systems Laboratory
Frank Battye \__/ <<<<< The Walter & Eliza Hall Institute
Dora Kaminaris ----------!!<<<<<<<< Victoria 3050, AUSTRALIA
Viki Lapatis /!!\ <<<<< ph: 61_3_9345 2540 fax: 61_3_9347 0852
o !! \ < in:: "facs_copy@wehi.edu.au"
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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