Search

Holographic Tomography (HT) is an emerging label-free technique for microscopic bioimaging applications, that allows reconstructing the three-dimensional (3D) refractive index (RI) distribution of biological specimens. Recently, an in-flow HT technique has been proposed in which multiple digital holograms are recorded at different viewing angles around the sample while it flows and rotates within a microfluidic channel. However, unlike conventional HT methods, there is no a priori information about cell 3D orientations, that are instead requested to perform any tomographic algorithm. Here we investigate a tracking-based rolling angles recovery method, showing robustness against the sample’s features. It is based on a phase images similarity metric recently demonstrated, that exploits the local contrast phase measurements to recognize a full cell rotation within the microfluidic channel. Hence, the orientations of the flowing cells are retrieved from their positions, which are in turn computed through the 3D holographic tracking. The performances of the rolling angles recovery method have been assessed both numerically, by simulating a 3D cell phantom, and experimentally, by reconstructing the 3D RI tomograms of two cancer cells. Both the numerical and the experimental analysis have been performed at different spatial resolutions. This rolling angles recovery method, not depending on the cell shapes, the RI contents, and the optical experimental conditions, could pave the way to the study of circulating tumor cells (CTCs) in the challenging tool of liquid biopsy.

Visualizing the intracellular dynamics of plant cells has been an open challenge for modern botany, agronomy and pharmacy. In this paper, we proposed an approach to improve the phase contrast during plant cell holographic imaging by cells' dehydration, and used this method to realize the observation of cytoplasmic circulation inside the living onion epithelial cell. The dehydration process can be seen as a sort of label-free contrast agent for better imaging biological processes. We have investigated live onion epidermal cells, observing their inner dynamics during long time recordings using a digital holographic microscopy system. For the experiments, an off-axis digital holography setup in transmission configuration with double spherical wave interference was used to record the digital holograms of onion cells. Then, we performed long-term time lapse holographic recordings of onion epidermal cells, and the results show that the intracellular tissue structure and the dynamic behavior of the cytoskeleton features and nuclei can be better exhibited via high-contrast phase imaging under cell dehydration conditions. In this case, the movements of intracellular filaments and the nucleus are observed via dynamical high-contrast phase imaging during the dehydration process. The experimental results clearly show the positive effect of dehydration process on intracellular imaging quality, and create the possibility to track the movement of plant organelles. In sum, thanks to the dehydration process of plant cells, holographic phase contrast enhancement imaging is realized.