Cytotoxicity quantification of nanoparticles is commonly performed by biochemical assays to evaluate their biocompatibility and safety. We explored quantitative phase imaging (QPI) with digital holographic microscopy (DHM) as a time-resolved in vitro assay to quantify effects caused by three different types of organic nanoparticles in development for medical use. Label-free proliferation quantification of native cell populations facilitates cytotoxicity testing in biomedical nanotechnology. Therefore, DHM quantitative phase images from measurements on nanomaterial and control agent incubated cells were acquired over 24 h, from which the temporal course of the cellular dry mass was calculated within the observed field of view. The impact of LipImage™ 815 lipidots^® nanoparticles, as well as empty and cabazitaxel-loaded poly(alkyl cyanoacrylate) nanoparticles on the dry mass development of four different cell lines (RAW 264.7, NIH-3T3, NRK-52E, and RLE-6TN), was observed vs. digitonin as cytotoxicity control and cells in culture medium. The acquired QPI data were compared to a colorimetric cell viability assay (WST-8) to explore the use of the DHM assay with standard biochemical analysis methods downstream. Our results show that QPI with DHM is highly suitable to identify harmful or low-toxic nanomaterials. The presented DHM assay can be implemented with commercial microscopes. The capability for imaging of native cells and the compatibility with common 96-well plates allows high-throughput systems and future embedding into existing experimental routines for in vitro cytotoxicity assessment.

The combination of micro synthetic structures with bacterial flagella motors represents an actual trend for the construction of self-propelled micro-robots. The development of methods for fabrication of these bacteria-based robots is a first crucial step towards the realization of functional miniature and autonomous moving robots. We present a novel scheme based on optical trapping to fabricate living micro-robots. By using holographic optical tweezers that allow three-dimensional manipulation in real time, we are able to arrange the building blocks that constitute the micro-robot in a defined way. We demonstrate exemplarily that our method enables the controlled assembly of living micro-robots consisting of a rod-shaped prokaryotic bacterium and a single elongated zeolite L crystal, which are used as model of the biological and abiotic components, respectively. We present different proof-of-principle approaches for the site-selective attachment of the bacteria on the particle surface. The propulsion of the optically assembled micro-robot demonstrates the potential of the proposed method as a powerful strategy for the fabrication of bio-hybrid micro-robots.

Optical tools for manipulation and trapping of micro- and nano-objects are a fundamental issue for many applications in nano- and biotechnology. This work reports on the use of one such method, known as photovoltaic optoelectronics tweezers, to orientate and organize cylindrical microcrystals, specifically elongated zeolite $L$, on the surface of Fe-doped ${\mathrm{LiNbO}}_3$ crystal plates. Patterns of aligned zeolites have been achieved through the forces and torques generated by the bulk photovoltaic effect. The alignment patterns with zeolites parallel or perpendicular to the substrate surface are highly dependent on the features of light distribution and crystal configuration. Moreover, dielectrophoretic chains of zeolites with lengths up to 100 μm have often been observed. The experimental results of zeolite trapping and alignment have been discussed and compared together with theoretical simulations of the evanescent photovoltaic electric field and the dielectrophoretic potential. They demonstrate the remarkable capabilities of the optoelectronic photovoltaic method to orientate and pattern anisotropic microcrystals. The combined action of patterning and alignment offers a unique tool to prepare functional nanostructures with potential applications in a variety of fields such as nonlinear optics or plasmonics.

The assessment of nanoparticle cytotoxicity is challenging due to the lack of customized and standardized guidelines for nanoparticle testing. Nanoparticles, with their unique properties, can interfere with biochemical test methods, so multiple tests are required to fully assess their cellular effects. For a more reliable and comprehensive assessment, it is therefore imperative to include methods in nanoparticle testing routines that are not affected by particles and allow for the efficient integration of additional molecular techniques into the workflow. Digital holographic microscopy (DHM), an interferometric variant of quantitative phase imaging (QPI), has been demonstrated as a promising method for the label-free assessment of the cytotoxic potential of nanoparticles. Due to minimal interactions with the sample, DHM allows for further downstream analyses. In this study, we investigated the capabilities of DHM in a multimodal approach to assess cytotoxicity by directly comparing DHM-detected effects on the same cell population with two downstream biochemical assays. Therefore, the dry mass increase in RAW 264.7 macrophages and NIH-3T3 fibroblast populations measured by quantitative DHM phase contrast after incubation with poly(alkyl cyanoacrylate) nanoparticles for 24 h was compared to the cytotoxic control digitonin, and cell culture medium control. Viability was then determined using a metabolic activity assay (WST-8). Moreover, to determine cell death, supernatants were analyzed for the release of the enzyme lactate dehydrogenase (LDH assay). In a comparative analysis, in which the average half-maximal effective concentration (EC₅₀) of the nanocarriers on the cells was determined, DHM was more sensitive to the effect of the nanoparticles on the used cell lines compared to the biochemical assays.

We report on a single capture approach for simultaneous incoherent bright field (BF) and laser-based quantitative phase imaging (QPI). Common-path digital holographic microscopy (DHM) is implemented in parallel with BF imaging within the optical path of a commercial optical microscope to achieve spatially multiplexed recording of white light images and digital off-axis holograms, which are subsequently numerically demultiplexed. The performance of the proposed multimodal concept is firstly determined by investigations on microspheres. Then, the application for label-free dual-mode QPI and BF imaging of living pancreatic tumor cells is demonstrated.

Microplastics are small plastic particles the size of less than 5 millimeters from cosmetics or results of abrasion and decomposition of plastic waste. The tremendous marine pollution by plastic particles and fibers and the increasing presence in the human environment from drinking water reservoirs to waste water demands for an environmental management and effective detection methods. The uptake of microplastics by living organisms may cause injuries of the gastrointestinal tract, trigger inflammation or cause cell toxicity by intrinsic particle properties or adsorbed pollutants. The urgent need for methods to identify microplastics in the environment, its sources of input and the risk of microplastic particles is the objective of the research project MicroPlastiCarrier. The project develops new tools for the optical detection and identification of microplastic particles from wastewater by a multiwavelength approach. The multiple labelfree optical toolbox is based on digital holographic microscopy using wavelengths from the visible to mid infrared. In order to monitor particle uptake minimally-invasively in living organisms and cellular specimens in a label-free manner, we applied high resolution optical coherence tomography (OCT) and multi-spectral digital holographic microscopy (DHM). In combination with microfluidics technologies as flow cytometry the project plans to identify particles based on size and their absorption and refraction index properties at several wavelengths. The technology should overcome the limitations of state of the art FT-IR.