http://confocal-microscopy-list.275.s1.nabble.com/Preamplifier-for-fast-point-scanning-tp7583344p7583409.html
thread could answer my question. Can someone describe the type of noise I
with megahertz few-pixel detectors. I assumed that the noise of each
settings. Is this true? Are there other important details I should know
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>> @Zdenek: A SNR of 3.5 or a little bit more might be enough for some
>> applications ... I was planning to measure the photon count per pixel
>> (using this method :
>>
http://labrigger.com/blog/2010/07/30/measuring-the-gain->> of-your-imaging-system/ ), but I always was busy with other things, so I
>> cannot give you numbers for my imaging system.
>>
>
> Hi all,
>
> Thanks to Labrigger for working on this important topic.
>
> However, I have read his analysis and think that the assumption that one
> can use this procedure to measure the number of photoelectrons (PE: i.e.,
> detected photons) created at the photocathode (PC) of the PMT may be an
> over-simplification.
>
> The analysis depends on the assumption that the only source of noise in
> the data recorded in the "image" of a flat white field is Poisson Noise
> associated with the small number of PEs produced at the photocathode. This
> might be true if PMTs were free from multiplicative noise but in fact
> Poisson Noise also affects every stage in the multiplication of a single PE
> after it leaves the PC. In the very unusual case that the voltage between
> the PC and the first dynode is 500-600 volts (and that this dynode has both
> the optimal shape and the best GaAs surface), the gain of this stage may be
> 25 +/-5 or 20% additional noise. More commonly, this gain will be closer
> to 4 +/-2 or 50% additional noise. More noise is added at each stage and
> even though these noise terms are not additive (they are combined as the
> sqrt of the sum of the squares), it is not at all uncommon for this process
> to double or even triple the variation present in the resulting signal
> beyond what one would expect from Poisson Noise applied only to the number
> of PE. Furthermore, this added noise will be somewhat larger if the system
> is working at a relatively high signal level because then the PMT will be
> turned down, the gain/stage correspondingly lower and the Poisson Noise
> proportionally higher.
>
> Offsetting this error to some extent is the finite bandwidth of the entire
> amplifier system (PMT plus the electronics between the final dynode and the
> ADC). This bandwidth is in general unknown but may be adjusted by the
> computer to more-or-less match what the computer estimates is needed to
> pass the finest optical details that the system can transmit on the basis
> of settings for wavelength, objective NA, zoom/pixel size, and even PMT
> setting (high PMT voltage implies a noisy signal that may benefit from the
> artificial, 1-dimensional smoothing that attends lower bandwidth).
>
> Clearly, because bandwidth limits the maximum excursion that can be
> transmitted between one pixel and its neighbour, it will tend to reduce the
> apparent noise present in the digitized signal. The magnitude of this
> clipping is unknown but may vary with the parameters mentioned above.
>
> This is relevant because, unlike the optical signal, the Poisson Noise
> signal that we are searching for shows no correlation between adjacent
> pixels. In particular, following the blog's suggestion of using a high zoom
> (to reduce fixed pattern noise) may cause the computer to limit the
> bandwidth more than using a lower zoom.
>
> Although, as noted above, because these two factors bias the results in
> opposite directions, their effects may cancel each other out to some
> extent. However, we need to know a lot more about how the components are
> actually operating before we can decide whether and to what extent this is
> true.
>
> The analysis also assumes that there is no fixed patterns noise in the
> image of a "flat white field" as might be caused, for instance, by field
> curvature, spherical aberration, vignetting, dust or other optical
> parameters that may change detected signal across the field of view. I
> note that many of these sources of non-Poisson Noise can be substantially
> reduced by recording two sequential frames and obtaining a measure of the
> noise by subtracting one from the other.
>
> For the analysis to work, it is also important to set the brightness
> control (DC - offset) so that zero signal corresponds to closely to zero
> intensity in the image memory.
>
> I should note that multiplicative noise ceases to be a factor in systems
> employing either hybrid PMT (where the first stage gain is about 10,000) or
> effective photon-counting (i.e. a photon counting where the recorded peak
> pixel signal is at least 10x smaller than the saturation count rate of the
> system as set by pulse-pileup.).
>
> One can avoid multiplcative noise by recording the data using a CCD (but
> NOT on an EM-CCD used with the electronic gain turned on) and the
> record-two-then-subtract approach can again be used to reduce inevitable
> fixed pattern noise. However, this sensor will probably work best when
> recording a fairly large signal (at least 10% of peak?) so that read noise
> will be relatively insignificant. And as above, the results will again be
> limited by the finite bandwidth of the FET amplifier between the read-node
> and the ADC. Finally, when using a CCD for quantitative measurements, it is
> particularly important to remember that they are usually set up so that
> zero light corresponds to 20-50 computer intensity units.
>
> The noise performance of sCMOS detectors is both non-Gaussian and depends
> strongly on the extent to which the internal pixel-by-pixel variations in
> gain and offset are detected and corrected. This will make their use for
> this type of measurement somewhat more difficult unless the signal levels
> are well away from the noise floor.
>
> Bottom line: Although the procedure may indeed give a useful benchmark
> that we might call the "effective gain" of the signal path, the measurement
> is subject to influence by a number of imaging parameters and will not
> really allow one to measure how many recorded-signal-intensity-units
> correspond to one PE.
>
> Jim Pawley
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