Abstract
Crucial benefits provided by partially coherent light microscopy such as improved spatial resolution, optical sectioning and speckle-noise suppression are exploited here to achieve 3D quantitative imaging: reconstruction of the object refractive index (RI). We present a partially coherent optical diffraction tomography technique (PC-ODT) that can be easily implemented in commercially available bright-field microscopes. We show that the high numerical apertures of the objective and condenser lenses, together with optical refocusing, are main issues for achieving fast and successful 3D RI reconstruction of weak objects. In particular, the optical refocusing is performed by a high-speed focus tunable lens mounted in front of the digital camera enabling compatibility with commercial microscopes. The technique is experimentally demonstrated on different examples: diatom cells (biosilica shells), polystyrene micro-spheres and blood cells. The results confirm the straightforward 3D-RI reconstruction of the samples providing valuable quantitative information for their analysis. Thus, the PC-ODT can be considered as an efficient and affordable alternative to coherent ODT which requires specially designed holographic microscopes.
Highlights
The three-dimensional (3D) reconstruction of the cell structure and its refractive index (RI) are required for disease diagnosis as well as for estimation of the cell dry mass and volume, etc
The main goal of our work is to demonstrate that the 3D RI distribution of weak objects, such as cells and micro-organisms, can be reconstructed by using a partially coherent optical diffraction tomography (PC-optical diffraction tomography (ODT)) technique developed for standard bright-field microscopes that exploits spatially low coherent illumination reaching a spatial resolution similar to the one obtained by the coherent ODT (C-ODT)
In contrast to previously reported works devoted to partially coherent microscopy imaging, here we experimentally demonstrate that the 3D refractive index distribution of the specimen can be straightforwardly reconstructed for the case of weakly absorbing samples by only measuring a single data set I r in the microscope under low spatially coherent illumination (S ∼ 1)
Summary
The three-dimensional (3D) reconstruction of the cell structure and its refractive index (RI) are required for disease diagnosis as well as for estimation of the cell dry mass and volume, etc. This is often referred to as beam rotation illumination and requires a temporal multiplexing approach to fill the maximum allowed 3D microscope aperture in order to reconstruct the 3D optical scattering potential, which is related to the refractive index of the sample This technique involves the acquisition and the computational processing of significant volume of data. The main goal of our work is to demonstrate that the 3D RI distribution of weak objects, such as cells and micro-organisms, can be reconstructed by using a PC-ODT technique developed for standard bright-field microscopes that exploits spatially low coherent illumination reaching a spatial resolution similar to the one obtained by the C-ODT.
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