Natacha Callens

Focus plane detection criteria in digital holography microscopy by amplitude analysis

We propose and test a focus plane determination method that computes the digital refocus distance of an object investigated by digital holographic microscopy working in transmission. For this purpose we analyze the integrated amplitude modulus as a function of the digital holographic reconstruction distance. It is shown that when the focus distance is reached, the integrated amplitude is minimum for pure amplitude object and maximum for pure phase object. After a theoretical analysis, the method is demonstrated on actual digital holograms for the refocusing of pure amplitude and of pure phase microscopic samples.

Digital holographic microscopy with reduced spatial coherence for three-dimensional particle flow analysis

We investigate the use of a digital holographic microscope working in partially coherent illumination to study in three dimensions a micrometer-size particle flow. The phenomenon under investigation rapidly varies in such a way that it is necessary to record, for every camera frame, the complete holographic information for further processing. For this purpose, we implement the Fourier-transform method for optical amplitude extraction. The suspension of particles is flowing in a split-flow lateral-transport thin separation cell that is usually used to separate the species by their sizes. Details of the optical implementation are provided. Examples of reconstructed images of different particle sizes are shown, and a particle-velocity measurement technique that is based on the blurred holographic image is exploited.

Extended focused imaging of a microparticle field with digital holographic microscopy

We present a numerical technique for extended focused imaging and three-dimensional analysis of a microparticle field observed in a digital holographic microscope working in transmission. The three-dimensional localization of objects is performed using the local focus plane determination method based on the integrated amplitude modulus. We apply the refocusing criterion locally for each pixel, using small overlapping windows, to obtain the depth map and a synthetic image in which all objects are refocused independent from their refocusing distance. A successful application of this technique in the analysis of the microgravity particle flow experiment is presented.

Hydrodynamic lift of vesicles under shear flow in microgravity

The dynamics of a vesicle suspension in a shear flow between parallel plates has been investigated under microgravity conditions, where vesicles are only submitted to hydrodynamic effects such as lift forces due to the presence of walls and drag forces. The temporal evolution of the spatial distribution of the vesicles has been recorded thanks to digital holographic microscopy, during parabolic flights and under normal gravity conditions. The collected data demonstrates that vesicles are pushed away from the walls with a lift velocity proportional to , where is the shear rate, R the vesicle radius and z its distance from the wall. This scaling as well as the dependence of the lift velocity upon the vesicle aspect ratio are consistent with the theoretical predictions by Olla (J. Phys. II 7 (1997) 1533).

Fast measurements of concentration profiles inside deformable objects in microflows with reduced spatial coherence digital holography

We investigate the use of a digital holographic microscope working with partially coherent spatial illumination to study concentration profiles inside confined deformable bodies flowing in microchannels. The studied phenomenon is rapidly changing in time and requires the recording of the complete holographic information for every frame. For this purpose, we implemented one of the classical methods of off-axis digital holography: the Fourier method. Digital holography allows one to numerically investigate a volume by refocusing the different planes of depth, allowing one to locate the objects under investigation in three dimensions. Furthermore, the phase is directly related to the refractive index, thus to the concentration inside the body. Based on simple symmetry assumptions, we present an original method for determining the concentration profiles inside deformable objects in microconfined flows. Details of the optical and numerical implementation, as well as exemplative experimental results are presented.

Particle Sorting In a Mini Step-Split-Flow Thin Channel: Influence of Hydrodynamic Shear on Transversal Migration

A mini splitterless-split-flow thin fractionation (SPLITT) device has been developed to achieve fast separations of micrometer-sized species. In this device, inlet and outlet steps have replaced the splitters, which are common to conventional SPLITT channels. By elimination of the splitters, it becomes straightforward to reduce channel dimensions while maintaining the classic method of fabrication. Reduced dimension channels allow high axial velocity at relatively low flow rate. These high axial velocities generate an enhancement of inertial lift forces and hydrodynamic shear-induced diffusion. Experiments carried out with particulate and biological species in a mini step-SPLITT channel demonstrate that these hydrodynamic effects yield highly enriched fractions of smaller species from binary mixtures.

Applications of digital holographic microscopes with partially spatial coherence sources

We implemented partially spatial coherent illuminations in digital holographic microscopes (DHM) working in transmission. The benefits gained with those sources are outlined. A major advantage is the drastic reduction of the speckle noise making it possible high image quality comparable to the best classical transmission microscopes. Several implementations of biomedical applications, where digital holography provides significant information, are described. With a rapid DHM permitting the analysis of dynamical phenomena, applications in microfluidics are also provided.

Digital Holographic Microscopy Working with a Partially Spatial Coherent Source

We investigate the use of partially spatial coherent illuminations for digital holographic microscopes (DHMs) working in transmission. Depending on the application requirements, the sources are made from a spatially filtered LED or from a decorrelated laser beam. The benefits gained with those sources are indicated. A major advantage is the drastic reduction of the speckle noise making possible high image quality and the proper emulation of phase contrast modes such as differential interference contrast (DIC) . For biomedical applications, the DHMs are coupled with fluorescence sources to achieve multimodal diagnostics. Several implementations of biomedical applications where digital holography is a significant improvement are described. With a fast DHM permitting the analysis of dynamical phenomena, several applications in fluid physics and biomedical applications are also provided.

Accurate three-dimensional detection of micro-particles by means of digital holographic microscopy

We present a numerical technique for refocusing and three-dimensional localization of micron-size particles observed in a digital holographic microscope working in transmission. We use Fourier method for the extraction of complex amplitude from the single exposition digital holograms. The three dimensional localization of objects is performed using the focus plane determination method based on the integrated amplitude modulus. We apply the refocusing criterion locally for each pixel, using small overlapping windows, in order to obtain a synthetic image in which all objects are refocused independent from their refocusing distance. We perform image segmentation and object detection using both the synthetic refocused image and the value of refocusing criterion, which allows us to obtain a high detection efficiency with very low number of false detections. While the lateral precision of localization is determined by the optical resolution of the setup, the vertical accuracy depends on the parameters of the digital holographic reconstruction. We improve the accuracy of vertical localization using an additional refining procedure in which each particle is treated separately. We analyze the robustness and accuracy of our approach and present its successful implementation in particle flow experiments.

Toward Prevention of Cowdriosis

Mass production of Ehrlichia ruminantium variants from different regions of sub‐Saharan Africa is one of the difficulties that must be overcome in producing a heartwater vaccine. Vaccine productivity can be limited by endogenous induction of interferon (IFN), which inhibits the propagation of Ehrlichia ruminantium (ER) in cell culture. Different kinds of endothelial cells, in which ER multiply efficiently, could be grown in a scalable way in VueLife Teflon bags on Cytodex 3 microcarriers where bead‐to‐bead transfer of cells occurs. The digital holographic microscope designed at the Université Libre de Bruxelles allows detection of the most appropriate time to harvest intracellular microorganisms for vaccine production.