I need a hand regarding photoactivable Ca/Voltage sensitive probes. I
for a Ca/Voltage sensitive probe with memory. I would like to find a
probe (ideally ratiometric) which ""freezes"" into its current state
In other words, I am interested in measuring the Ca concentration at a
point in time and from many location in the tissue (in particular at
several dephs in the tissue).
Because the number of cells from which I want to do the measurement
is high, I can not use a time changing probe. Ideally, I would like to
have a "freezing" probe
which retains a particular conformation (which must be Ca or voltage
illumination with the activation wavelength. After illumination, I would
scan the whole
tissue measuring the Ca concentration that was in the cell at the point
in time of interest.
There may be an alternative to having such a complex probe. May be
In another variation of the experiment, I would like to have an in-vitro
preparation and apply
the "freezing" Ca sensitive probe. After some time, I would activate the
Ca probe which would
freeze into one of two fluorescense states (A which emits wavelength A
or B which emits wavelength
B). The probability of freezeing in state A or state B should be
dependent on Ca concentration (or voltage).
After the activation flash, I would fixate the tissue (with a protocol
which conserves fluorescense
from the probe) and microtome-slice the tissue. Then, I would extract
3D info about the Ca concentration
at the time of the activation flash.
I will appreciate any suggestions and apologies for this long email.
Enric T. Claverol
University of Southampton