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An analogue-to-digital unit is a unit of a digitized photodetector output after electronic conversion. Its relation to collected Electrons is set by the detector conversion gain; an additive electronic offset must be removed before interpreting a count as a light signal.

For stable positive detector pixel means $N_i$ and uniform detector conversion gain $g$, high-count independent Poisson distribution noise gives $\mathrm{Var}(A_i/B_i)\simeq 2/(g{N}_i)$. Thus substituting a global mean in the equal-signal formula yields $g_{\mathrm{naive}}=g/[⟨N_i⟩⟨1/N_i⟩]\le g$, by the Jensen inequality. Equality holds for uniform means. The flat-field correction pattern cancels from the ratio mean but not from the signal-dependent noise. Local signal bins, or a variance fit after normalization of differences by the square root of their local mean, avoid this leading bias.

Detector conversion gain is the number of collected Electrons per analogue-to-digital unit. With independent Poisson distribution photoelectron counts, the shot-noise variance in count units is the mean count divided by the gain. Detected-photon and Electron conventions agree only when each detected photon contributes one Electron.

Let $k=m/2$. After offset subtraction, each summed image has mean $N$ analogue-to-digital units per detector pixel. For a constant detector conversion gain $g$ and independent Poisson distribution detected counts, $\mathrm{Var}(A)=\mathrm{Var}(B)=N/g$: summing $k$ independent frames adds both their means and variances. Here $N$ is the mean of the sum, not one frame.

Write $A=N+a$, $B=N+b$. At high counts, the delta method gives $A/B\simeq 1+a/N-b/N$. Independence therefore givesThus the formula is a high-signal gain estimation from flat-field ratios, not an exact identity for a ratio of random variables. A Poisson distribution denominator can even be zero at low counts. The exposures must be linear and unsaturated, offsets removed, and read noise negligible. Being close to saturation increases the signal but does not establish linearity; clipping or charge-induced correlations biases a variance estimate. A large ensemble of suitable detector pixels supplies the measured standard deviation. In normal photodetector terminology $g$ is collected Electrons per ADU, consistent with detected photons here only under the stated unit-yield convention; see the instrument gain definition.

If spatial illumination is stable between the two sums, it acts just like the sensitivity factor: the mean ratio remains one but each detector pixel has a different shot-noise variance. A whole-image standard deviation is disproportionately influenced by faint detector pixels, so substituting a global arithmetic mean signal generally underestimates detector conversion gain. If the illumination pattern differs between the sums, their noise-free ratio is not constant; its structure adds apparent variance and causes a further systematic error.

Use locally uniform signal bins or subregions, remove offsets, mask defective or saturated detector pixels, and propagate the local shot and read variances. In the stable-pattern case a useful alternative to pooling raw ratios is the normalized differenceAt high signal and negligible read noise, treating the denominator at its mean gives $\mathrm{Var}(U_i)\simeq (2N_i/g)/(2N_i)=1/g$, independent of the illumination. Thus a variance fit or $g\simeq 1/\mathrm{Var}(U)$ can use a spatially varying flat without the arithmetic/harmonic mismatch. Measured noisy denominators cause higher-order corrections, so independent or smoothed signal estimates can be preferable. Fit and subtract a genuine changing illumination pattern before noise analysis, using sufficient smoothing not to fit away the random noise. Arbitrary flat-field normalization without propagating its noise does not by itself recover the original gain.