Ion ratio imaging is a relatively new technique
for measuring either absolute or relative changes in ion
concentration within living cells. This is accomplished
by measuring the fluorescence emission of special dyes
which have been designed to change their spectral
properties or emission intensities upon binding to the
ion of interest.
RatioImg is a suite of Igor Pro 3.13 procedures for
processing and analyzing data from ratiometric imaging
experiments (i.e., A and B wavelength measurements of
multiple ROIs); it was originally developed for use with
the sodium indicator dye SBFI. RatioImg can handle
single-wavelength or already-ratioed data; it can perform
background subtractions and can correct ratio values
based on in situ fluorescence measurements using
ionophores and known concentrations of the ion in the
perfusion bath. For single-wavelength dyes, it can
calculate and display F/Fo or delta_F/F. Data from each
cell (ROI) can be browsed and displayed in a graph
window, graphs can be copied to a notebook and annotated,
and a variety of measurements can be performed.
Detailed on-line help is included along with complete
documentation in pdf format. There are also two sample
data files for use with the documentation's tutorial
section and an experiment file showing examples of
annotation and measurement.
The Macintosh or Windows versions of RatioImg can be
downloaded using one of the following links:
We have tested RatioImg on a Mac PPC 6400/200 with OS
8.1, and on a Dell Optiplex GXM 5166 with Win95. There
are slight differences between the Mac (.sit) and Windows
(.zip) archives due to mostly cosmetic OS differences.