[Cytometry] Cytometry] seeking advice with measuring Calcium using Fura-3 and Fluo3 or 4?
g.warnes at qmul.ac.uk
Wed Feb 4 04:08:55 EST 2009
I have a protocol on my website for Fluo-4, see link
normally use Indo-1 instead of Fura-2 on flow cytometers as this excites
with UV and emits at about 400nm and over 500nm enabling a ratiometric
measurement see the link
Fura-2 is normally used on microscopes as this dye is excited with two
wavelengths which are between 300 & 400nm so you could try using UV and the
405nm line to excite the Fura-2 (could not find Fura-3 in the Molecular
probes hand book) but I'm not sure this would work as nicely as Indo-1 does.
Are you comparing the FACS calcium with imaging with Fura-2 I was asked to
do this once but then my confocal is set up to run Indo-1 so we switched
I normally do my Calcium experiments at RT as they work fine in most cases
if you need to use 37C then I put a small beaker of 37C water on the probe
at the same time as the tube and just replace the water after every run.
This approach is a bit difficult after you have added the agonist to the
tube and replacing the tube quickly on the machine so you might want to use
the Vantage setup.
Best of luck with this experiment
Dr Gary Warnes
Flow Cytometry Manager
Flow Cytometry Core Facility
Institute of Cell & Molecular Science
Barts and The London School of Medicine and Dentistry
4 Newark Street
Fax No. 0207-882-2180
I was wondering if someone could help me with a protocol for measuring
intracellular Ca++ in murine T cells using the probes Fura-3 and Fluo-4.
I didn't find more than a couple of citations of the 2006 paper by Sheree
Bailey and MacArdle, and I was wondering if this assay is routinely being
used on a FACS Canto II or LSR II. Is there an advantage to using a FACS
Vantage's temperature controlled sample holder instead of the benchtop
Andrew Herman, Ph.D.
University of Bristol
Department of Cellular & Molecular Medicine Director, Flow Cytometry
Facility School of Medical Sciences University Walk Bristol BS8 1TD UK
Mobile: 079 6782 9670
A.Herman at bristol.ac.uk
More information about the Cytometry