CONFOCOR
3
Carl Zeiss
Photon Counting Imaging with the ConfoCor 3
ConfoCor 3
136 M60-1-0025
e
02/2010
9.14.2.1
The Analog (Grey Value) Display
In this display, photons are transformed into grey values, which are displayed in discrete intervals in the
histograms. Intervals can be calculated by:
Interval = (pixel period x frequency) / (2n-1) = (pixel period x CRmax/digital gain) / (2n-1),
where n = 8 for an 8 bit image and n = 12 for a 12 bit image (see Fig. 155 for an example).
Fig. 155
Analog display in photon counting image. Settings were digital gain 1, scanning speed 8
corresponding to a pixel period or pixel dwelling time 2.56 µs, 8 bit image and APD detectors
Example:
With the APD as the detector, software would limit the maximum count rate to 4 Mhz / 1 = 4 photons / µs.
At scanning speed 8 with a pixel dwelling time of 2.56 µs one can obtain a maximum of 4 x 2.56 = 10.24
photons. Since photons are integer, this value must be rounded down to 10 photons maximum / pixel.
The spacing interval is calculated for the 8 bit image as 255/10=25.5. This value is rounded down to the
next fifth value, which is 25.
So 1 photon will correspond to a grey value 25, n photons to a grey value n x 25, with a maximum
n of 10).
Grey value 0 is reserved for 0 photons, 255 for any overflow. In this way, the histogram is stretched to
cover as much as possible of the available grey scale. Therefore, even at lesser or greater photon counts,
the image will maintain its brightness.
In the
Status Area
the grey value of the pixel to which the mouse points in the image is displayed. A
grey value of 75 would mean 75/25=3 photons in the example with speed 8.
Analog images can be used for all further processing in the LSM software like mean of ROI
function, statistics etc.
