We present a stimulated emission depletion (STED) microscope that provides 3‐D super resolution by simultaneous depletion using beams with both a helical phase profile for enhanced lateral resolution and an annular phase profile to enhance axial resolution. The 3‐D depletion point spread function is realised using a single spatial light modulator that can also be programmed to compensate for aberrations in the microscope and the sample. We apply it to demonstrate the first 3‐D super‐resolved imaging of an immunological synapse between a Natural Killer cell and its target cell. (© 2014 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)

TIRF and STORM microscopy are super-resolving fluorescence imaging modalities for which current implementations on standard microscopes can present significant complexity and cost. We present a straightforward and low-cost approach to implement STORM and TIRF taking advantage of multimode optical fibres and multimode diode lasers to provide the required excitation light. Combined with open source software and relatively simple protocols to prepare samples for STORM, including the use of Vectashield for non-TIRF imaging, this approach enables TIRF and STORM imaging of cells labelled with appropriate dyes or expressing suitable fluorescent proteins to become widely accessible at low cost.