UV (280 nm) excitation in flow for lanthanides?

Robert C. Leif, Ph.D. rleif at rleif.com
Mon Apr 17 17:22:08 EST 2000


From: Bob Leif
To: cyto-inbox

My original suggestion (1) to measure Trp fluorescence in flow was to use a
relatively inexpensive flashlamp. The lamp could be triggered by either low
angle light scatter or, as originally suggested, the DC Coulter Effect. If
the cells are flowed slowly, about 500 microseconds observation period, and
the flash is upstream of the optical detection field, you might obtain a
decent signal. The use of deuterium oxide should also be considered. if you
are successful, please let me know. I have other compounds that could be
used.

(1) R. C. Leif; “A Proposal for an Automated Multiparameter Analyzer for
Cells (AMAC)”. Automated Cell Identification and Sorting, Edited by G. L.
Wied and G. F. Bahr, Academic Press, New York, pp. 131-159 (1970).

-----Original Message-----
From: Howard Shapiro [mailto:hms at shapirolab.com]
Sent: Friday, April 14, 2000 3:14 PM
To: cyto-inbox
Subject: Re: UV (280 nm) excitation in flow for lanthanides?



Dane Wittrup writes:


>Does anybody have their flow instrument set up for excitation from 280-300
>nm?  We'd like to excite a lanthanide bound in calmodulin, using Trp as the
>donor to the ion.

There aren't a lot of people who have lasers that will do those deep UV
lines - the only publication in this area which comes to mind was from the
Jovins in Goettingen in the late 1970's, looking at Trp fluorescence in
proteins.

However, if you're really interested and have the laser (which would
presumably be on an optical bench) and optical filters, I could bring over
the rest of a cytometer.  You might also be able to scrounge the
do-it-yourself cytometer parts from Penny Chisholm's lab, which is even
closer to you.

Of course, the real problem might be the lanthanide lifetime; as I recall,
this runs into many microseconds, while the typical dwell time in the beam
of a cytometer is only a few microseconds.  Call if you're interested
(617-783-8392 or 617-965-6044).

-Howard






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