I've tried observing cells using luciferase activity in the cytometer,
without success.
Luciferin requires ATP and molecular oxygen to drive the reaction, in the
presence of Mg2+. The difficulty I had (I think) was getting the
substrate (luciferin), and ATP, into the cells without damaging them. Even
using a caged luciferin, which is cell permeant and activated
intracellularly, didn't work. If anybody's got past that stage or wants
to discuss further off line, I'd be very keen to hear from them.
Simon
Simon QJ Rice
GlaxoSmithKline R&D
Harlow, UK
"Richard Haugland" <richard.haugland@probes.com>
05-Dec-2002 03:02
To: "Cytometry Mailing List"
cc:
Subject: Re: luciferase essay using flow cytometer?
Interesting question. The chemiluminescence peak of luciferin is at
about 560 nm, which is a bit shorter wavelength than phycoerythrin
emssion. Of course, it does not need any excitation to get the emission.
But then, I don't know if the rate of emission will be fast enough to
get enough photons during the transit time that you will be able to
detect the signal. But then S/N should be great because all photons
should be "real." I don't know what technical difficulties there may be
doing sorting with the laser turned off but hope it works.
Nan Jiang wrote:
>Dear all:
>
>I am wondering if anybody ever sorted cells infected with luciferase
>plasmid. Or is there a particular wavelengh we can use to detect
>luciferase activity in cells and sort them based on that.
>
>Thanks in advance.
>
>Nan Jiang
>
>Department of Internal Medicine
>Cardiology Division
>UT Southwestern Medical center at Dallas
>6000 Harry Hines Blvd.
>Dallas, TX 75390-8573
>(214) 648-1175
>(214) 648-1181
>
>
>
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