We present a combination of nonlinear microscopy and optical trapping applied to three-dimensional imaging and manipulation of intracellular structures in living cells. We use Titanium-sapphire laser pulses for nonlinear microscopy of the nuclear envelope and the microtubules marked with green fluorescent protein in fission yeast. The same laser source is also used to trap small lipid granules naturally present in the cell. The trapped granule is used as a handle to exert a pushing force on the cell nucleus. The granule is moved in a raster-scanning fashion to cover the area of the nucleus and hence displace the nucleus away from its normal position in the center of the cell. Such indirect manipulations of an organelle (e.g., nucleus) can be useful when direct trapping of the chosen organelle is disadvantageous or inefficient. We show that nonlinear microscopy and optical manipulation can be performed without substantial damage or heating of the cell. We present this method as an important tool in cell biology for manipulation of specific structures, as an alternative to genetic and biochemical methods. This technique can be applied to several fundamental problems in cell biology, including the mechanism of nuclear positioning and the spatial coordination of nuclear and cell division.
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