How to do method Validation in microscopy?

Calibration and Validation is an important activity in Microscopy Particle size and shape Analysis. Calibration of the software is done by calibrating the Micrometer scale or slide called as Calibration slide. Calibration is important because we guide the software on how to convert pixels to micrometres and because of that, the sample images with particles will easily provide and identified its particles with accurate morphological details.
Validation is done by a certified predefined diameter of circles or predefined shapes; the Microscopic software system gives the result of certified predefined circles and shapes and then compared them with the actual size and shape. If it is within the acceptable limit, the software system is validated successfully.