FN ISI Export Format VR 1.0 PT J TI Nanometry of bio-molecular complexes using PSF reconstruction in SMI microscopy AF Birk, Udo Baddeley, David Mathée, Hans Wagner, Christian Cremer, Christoph AU Birk, U Baddeley, D Mathée, H Wagner, C Cremer, C AB Spatially Modulated Illumination Microscopy (SMI) in combination with reference objects of known size to reconstruct the Point Spread Function (PSF) is applied to determine the size of individual fluorescent objects. SMI employs a structured illumination to obtain information about the specimen far beyond the conventional resolution limit. Size determination is based on small perturbations of the intensity modulation by the fluorescent objects. The SMI-PSF is used to derive the object size from these perturbations. Reference objects with known dye distribution are added to the sample to reconstruct the PSF. We present data from measurements on fluorescent microspheres with diameters between 40 and 200 nm using an excitation wavelength of 488 nm. Applications of gene expression analysis, transcription factories analysis, protein cluster analysis, and gene compaction analysis prepared on conventional microscope slides are shown. The size values obtained are about two orders of magnitude smaller compared to the extension of the illumination or observation volume, respectively, in a confocal laser scanning microscope using a high numerical aperture objective lens. ER