Disaggregation of colonies of Microcystis (Cyanobacteria) into smaller aggregates or single cells is a useful process for improving the accuracy and precision of cell abundance estimates in natural populations. An image analysis system was therefore used to assess the efficiency of two techniques, heating/vortexing and grinding, for disaggregating fixed and live colonies. Eight environmental samples, consisting of either colonial or non-colonial strains, were tested in time course experiments using each technique by analysing the number and total area of the objects (i.e., cells, clumps of cells, colonies) acquired. The results show that i) image analysis is an efficient tool to compare the efficiency of disaggregation techniques on colonies of cyanobacteria, ii) the grinding technique is more efficient in disaggregating all types of Microcystis colony, without significant loss, and iii) the heating technique does not work for all types of colonies, in particular, compact colonies fixed with Lugol’s iodine solution.