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Dear all,
First of all, hello everyone.
I am new to the mailling list and the microscopy world.
I'm a portughese student and am currently doing my master thesis at the
Instituto Gulbenkian de Ciência under the guidance of Nuno Moreno.
We are trying to measure the light capturing efficiency of a confocal
microscope but aren't sure of the methodology and were hoping you could
shed some light on it.
The efficiency we want to measure is the fraction of photons emitted from a
light source that is detected in the final image, being: efficiency= np/ne,
where np is the number of detected photons and ne is the photons emitted by
a calibrated light source(LED).
To calculate np we assumed there was no amplification noise, and used the
mean of the image M and it's variance V.
M=np*g and V=np*g^2.
Being g the average pixel intensity resulting from the detection of one photon,
which is proportional to the gain of the PMT and subsequent elecronics. And
np the average detected photons per pixel.
Our doubt is in the calculation of ne.
Is each pixel a representation of all the light that reaches the lens during the
pixel dwell time?
If so, ne should be the number of photons that reach the lens during the dwell
time.
If not, if each pixel represents only a portion of the light that reaches the lens
during the dwell time, than ne should be a fraction of the number of photons
that reach the lens during the dwell time.
I have searched, but I am unable to find any kind of tutorial to help me
understand this concept. If somebody could point me in the direction of a
good source to understand how this should be done, I would be very grateful.
Many thanks
Francisco Henriques
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