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International audience; This chapter starts with raw images and extracts objects, analyses them further to yield quantitative information from them. It guesses from the initial musing that doing quantification involves mastering computers and being familiar with some mathematical concepts. Quantification of image data can be seen as a subset of image analysis techniques that provides answers in the shape of numbers, extracted from images. An understanding of the basic mathematical principles which they rely on is essential to performing scientifically relevant quantitation. Most of the image analysis tools will have some means of carrying out the calculations and yield this quantifiable information. There is a whole world of quantifiable information to be found in time‐lapse images, fuelled for instance by single‐particle tracking algorithms, optic‐flow techniques and all image analysis techniques able to harness time. Also, some information can be extracted from bulk images, without relying on object extraction.

International audience; This chapter surveys basic segmentation techniques and point to tools that make them accessible in practical situations. Segmentation is very often the first step when doing quantification. Other image analysis techniques, such as pixel‐based co‐localisation, optical flow, etc. are not based on segmentation, yet offer useful information condensation that can lead to a proper scientific conclusion. The chapter focuses on the list of integers that comprises an image. Automatic counting of objects is almost always done on images where objects appear bright over a dark background, ideally flat. The set of connected pixels with a shared identity forms an object, from which a contour can be extracted and quantification performed. The chapter focuses on what quantifiable information is required to reach a scientific conclusion, pick an analysing technique that can yield it, find or write a software tool that implements it, and finally acquire images that can be analysed with such a tool.