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We report on the design and construction of a laser scanning photothermal microscope and present images of gold nanoparticles of size as small as 5 nm. Laser scanning method allows fast image acquisition at 80 μs pixel dwell time so that a 500 × 500 pixel image is acquired in 20 s. Photothermal imaging at fast time scales can have potential applications in variety of fields including tracking of biomolecular transport processes. [ABSTRACT FROM AUTHOR]

Nuclear pore complexes in the nuclear membrane act as the sole gateway of transport of molecules from the cytoplasm to the nucleus and vice versa. Studies on biomolecular transport through nuclear membranes provide vital data on the nuclear pore complexes. In this work, we use fluorescein isothiocyanate-labeled dextran molecules as a model system and study the passive nuclear import of biomolecules through nuclear pore complexes in digitonin-permeabilized HeLa cells. Experiments are carried out under transient conditions in the time lapse imaging scheme using an in-house constructed confocal laser scanning microscope. Transport rates of dextran molecules having molecular weights of 4-70 kDa corresponding to Stokes radius of 1.4-6 nm are determined. Analyzing the permeability of the nuclear membrane for different sizes the effective pore radius of HeLa cell nuclear membrane is determined to be 5.3 nm, much larger than the value reported earlier using proteins as probe molecules. The range of values reported for the nuclear pore radius suggest that they may not be rigid structures and it is quite probable that the effective pore size of nuclear pore complexes is critically dependent on the probe molecules and on the environmental factors. [ABSTRACT FROM AUTHOR]