Fluorescent observation of cells generally suffers from the limited axial resolution
due to the elongated point spread function of the microscope optics. Consequently,
three-dimensional imaging results in axial resolution that is several times worse
than the transversal. The optical solutions to this problem usually require complicated
optics and extreme spatial stability. A straightforward way to eliminate anisotropic
resolution is to fuse images recorded from multiple viewing directions achieved mostly
by the mechanical rotation of the entire sample. In the presented approach, multiview
imaging of single cells is implemented by rotating them around an axis perpendicular
to the optical axis by means of holographic optical tweezers. For this, the cells
are indirectly trapped and manipulated with special microtools made with two-photon
polymerization. The cell is firmly attached to the microtool and is precisely manipulated
with 6 degrees of freedom. The total control over the cells' position allows for its
multiview fluorescence imaging from arbitrarily selected directions. The image stacks
obtained this way are combined into one 3D image array with a multiview image processing
pipeline resulting in isotropic optical resolution that approaches the lateral diffraction
limit. The presented tool and manipulation scheme can be readily applied in various
microscope platforms.
