Axial Tomography in Single Cell Fluorescence Microscopy

Axial tomography in combination with various techniques of 3D microscopy including confocal laser scanning microscopy, structured illumination microscopy or light sheet based fluorescence microscopy improves visualization of single living cells or small cell clusters in three dimensions. After adaptation of an innovative device for sample rotation, single cells embedded in agarose and located within micro-capillaries are observed from different sides. Thus, z-stacks of 3D samples can be recorded in different directions, providing additional information, since cells or organelles which appear superimposed in one direction, may be well resolved in another one. Furthermore, since lateral resolution is better than axial resolution by at least a factor 2, an improved effective 3D resolution is achieved upon sample rotation. Axial tomography in combination with Confocal Laser Scanning Microscopy (C-LSM) represented a step towards high resolution microscopy of single cells. The uptake of fluorescent dyes and nano-particles was well documented in the sub-micrometer range. The technique appeared also appropriate to measure nuclear architecture upon uptake of the anticancer drug doxorubicin.