UV excitable dyes for surface phenotyping - ELF-97?

Marty Bigos mbigos at gladstone.ucsf.edu
Wed Mar 13 18:31:09 EST 2002


>Hi all,
>	We are in the process of getting some extra colours working
>on our new FACSVantage SE and are trying to find a dye we can excite
>with the third laser (a mixed gas argon and krypton (Coherent Innova
>70 Spectrum UV)).
>
>I have read the Molecular Probes literatures and the publication by
>Telford et al (Cytometry 32:117-125 (2001)) about the use of the
>ELF-97 enzyme substrate system and was wondering if anyone has any
>"real-world" experience using it with surface antigens? I am
>particularly concerned about the 30min incubation at RT that is
>required. What does this do to live cells? (the MP literature
>suggests using fixed cells but the Cytometry paper seems to use live
>cells). What about the labelling of other surface markers?
>
>Any other suggestions for alternative dyes that we can use with this laser?

The original publications using cascade blue excited it with the UV
from an argon laser. However, because of high autofluorescence, it
was useful mainly as a second step with "bright" antibodies. We have
used cascade blue conjugates with 407 nm krypton excitation; they are
usable, but not great. Also, Molecular Probes makes Alexa Fluor 350
conjugated as a second step reagent, which might be useful as well.
We have no experience with it here.

Marty


>
>Thanks,
>
>Adrian Smith
>
>--
>______________________________________________
>Adrian Smith (Research Officer)	T CELL BIOLOGY GROUP
>Centenary Institute of Cancer Medicine & Cell Biology
>Locked Bag No.6 Newtown, NSW 2042 AUSTRALIA.
>Ph: (02) 9565-6198 Fax: (02)-9565-6103

--
Marty Bigos
Director, Flow Core
Gladstone Institute of Virology and Immunology
Building 3 SFGH Rm 509
415-695-3832



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