Area from a calibrated pixel count ($A=N\Delta x\Delta y$)

Area from a calibrated pixel count assigns an area to a selected collection of complete image pixels. For $N$ selected pixels of calibrated widths $\Delta x$ and $\Delta y$,This rectangular-cell model permits unequal horizontal and vertical spacing. In ImageJ, area is reported in square pixels or calibrated square units according to the spatial calibration and measurement settings.^[ref]

For an original threshold example, let four stored gray values be 40, 90, 130, 200. A rule selecting values at least 128 retains two pixels. If $\Delta x=\Delta y=0.5\mu \mathrm{m}$,At $\Delta x=0.5\mu \mathrm{m}$ and $\Delta y=0.8\mu \mathrm{m}$, the same pixel selection instead represents $0.80\mu {\mathrm{m}}^2$. Pixel count alone is not an area in physical units.

For a binary selection mask $M_{ij}\in \{0,1\}$, the count is $N={\sum }_{ij}{M}_{ij}$. A fractional-coverage model would instead use fractions $0\le f_{ij}\le 1$ and $A=\Delta x\Delta y{\sum }_{ij}{f}_{ij}$; the two rules need not agree at boundaries. A region of interest and an image thresholding rule also select pixels for different reasons. State the selection and calibration before interpreting the result as an estimate of a specimen's geometric area. The numbers above are stipulated image data, not a physical specimen measurement.