Nanomanipulation in the context of biology is a well discussed and reviewed topic in the scientific community in the last decades.1 The ability to control an arm at the nanoscale level has shown promise in material sciences and biology.2,3 A simple way to achieve this small scale manipulation in a lab is to use an atomic force microscope (AFM), a derivative from scanning probe microscopes (SPMs).4 AFM measures the deflection of a cantilever, which is caused by the interaction with the surface of the sample and a sharp tip that extends from the free end of the cantilever. This deflection characterizes the distance from the sample and force felt by the tip. The SPM methods initially worked on conductive samples and vacuum conditions giving resolution at the subangstroms scale, but AFM gained its popularity and demand because of its ability to work also on insulating materials and in liquid environments; especially in physiological conditions.5 Commercial AFMs are now designed to work in a dry environment for material sciences applications and wet environments for investigating biological systems. AFM is an established technique in the study of cellular morphology and local stiffness, understanding protein folding mechanisms, checking model membranes stability, etc.6–9 As pointed out before, the AFM cantilever with the help of its extended stylus reaches the sample and precisely calculates the surface height. However, if more force is applied on this stylus, the stylus mechanically interacts with the sample. The spatial precision of this interaction is defined by the AFM tip size that could be tens of nanometres, but this manipulation is limited by its nonspecific nature in identifying the sample10,11 or selecting a target for manipulation. An optical microscope coupled to an AFM can recognize and target an area of interest with the help of specific identification markers like fluorescence tags. In the past years, superresolution (SR) fluorescence microscopes have taken the place of conventional fluorescence microscopes like confocal and widefield microscopes. The SR microscopy methods visualize the fluorescence below the classical optical diffraction limit () and reach nanometre level lateral resolution. An example of SR microscope techniques such as stimulated emission depletion (STED)12 microscope is an improvement over the confocal microscopes. This article demonstrates how to build an STED microscope from a commercial multiphoton microscope with its complete building blueprint.