Calibrating photon counts from a single image

in: Nature Communications (2018)
Heintzmann, Rainer; Relich, Peter K.; Nieuwenhuizen, Robert P.J.; Lidke, Keith A.; Rieger, Bernd
Most image capturing devices do not directly report the number of detected photons, but a value proportional to the photoelectron charge produced in a photomultiplier tube or collected in a camera pixel. In order to establish the photon count, the gain of the device must be measured, typically by recording tens of calibration images and exploiting the linear relationship between mean intensity and its variance [vanVliet1998]. Here we propose and evaluate a method that obtains the gain from a single acquired image by quantifying out-of-band information. As noise is not limited to the cut-off frequency of the optical transfer function (OTF), estimation of the out-of-band energy relative to the total energy enables computation of the gain. We show on simulation and experimental data that this much simpler procedure, which can be retroactively applied to any image, is comparable in precision to traditional gain calibration procedures.

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