Bi-plane Calibration in Super-resolution Microscopy

In the localisation stage, the two stacks are processed separately. Local maxima are assumed to originate from fluorophore beads, and a threshold value is then used to keep the prominent ones only [2]. Each one of these local maxima is then fitted with a PSF model, yielding the lateral and the axial coordinates of each fluorophore bead. The fitting error is further compared with a threshold value for obtaining reliable localisation results. The algorithm fits the local maxima with the Gibson and Lanni PSF [3]. This model considers aberrations that are due to refractive indices mismatches at the sample cover slip immersion interfaces, as well as for the thickness of these layers. In particular, axial stage displacements correspond to different values of the immersion layer thickness. The output of the localisation stage is two lists of 3D beads locations.