Application of STED imaging for chromatin studies

Georgij Kostiuk, Jonas Bucevičius, Rūta Gerasimaitė and Gražvydas Lukinavičius

Chromatin is the information center of a cell. It comprises proteins and nucleic acids that form a highly complex and dynamic structure within the nucleus. Its multiple organization levels span from micrometre to nanometre scale. For many years, the lower levels of chromatin organization have been beyond the resolution limit of fluorescent microscopy, thus impeding research on nucleus architecture, transcription, translation and DNA repair. Recent development in super-resolution fluorescence microscopy enables us to more easily observe objects at the nanometre scale and allows the study of complex cellular structures at unprecedented detail. This review focuses on the application of stimulated emission depletion microscopy for imaging two main components of the chromatin-DNA and the proteins interacting with it.

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Resonant dielectric metasurfaces: active tuning and nonlinear effects

Chengjun Zou, Jürgen Sautter, Frank Setzpfandt and Isabelle Staude

Resonant dielectric metasurfaces were extensively studied in the linear and static regime of operation, targeting mainly wavefront shaping, polarization control and spectral filtering applications. Recently, an increasing amount of research focused on active tuning and nonlinear effects of these metasurfaces, unveiling their potential for novel nonlinear and reconfigurable optical devices. These may find many applications in imaging systems, compact adaptive optical systems, beam steering, holographic displays, and quantum optics, to just name a few. This review provides an overview of the recent progress in this field. Following a general introduction to resonant dielectric metasurfaces, the current state-of-the-art regarding the enhancement and tailoring of nonlinear frequency conversion processes using such metasurfaces is discussed. Next, we review different approaches to realize tunable dielectric metasurfaces, including ultrafast all-optical switching of the metasurface response. Finally, future directions and possible applications of nonlinear and tunable dielectric metasurfaces will be outlined.

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Colloidal rods in optical potential energy landscapes

Joshua L. Abbott, James A. Spiers, Yongxiang Gao, Dirk G A L Aarts and Roel P A Dullens

We study the static and dynamic behaviour of colloidal rods in an optical potential energy landscape. We explore the stable states of a colloidal rod in a single optical trap close to a flat wall. Here two metastable states are observed, horizontal and vertical, both of which experience a parabolic potential energy landscape. Next we place a colloidal rod into a one-dimensional sinusoidal optical potential energy landscape and introduce a constant driving velocity. When driven below the critical velocity, the particle is confined to a single potential energy minimum of the optical landscape and the equilibrium position of a particle is investigated. The equilibrium position of a rod is found to vary substantially from that of a sphere due to the drag coefficient of a rod, which is highly dependent on its proximity to an optical trap. The driving velocity is increased to enable the particle to traverse the periodic landscape and above the critical velocity, the average particle velocity increases as the square root of the driving velocity. When introducing oscillations to the driving velocity we observe dynamic mode locking and characterise the nature of synchronised motion of the rod-like particles.

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Physical Probing of Cells

Florian Rehfeldt and Christoph F Schmidt

In the last two decades, it has become evident that the mechanical properties of the microenvironment of biological cells are as important as traditional biochemical cues for the control of cellular behavior and fate. The field of cell and matrix mechanics is quickly growing and so is the development of the experimental approaches used to study active and passive mechanical properties of cells and their surroundings. Within this topical review we will provide a brief overview, on the one hand, over how cellular mechanics can be probed physically, how different geometries allow access to different cellular properties, and, on the other hand, how forces are generated in cells and transmitted to the extracellular environment. We will describe the following experimental techniques: atomic force microscopy, traction force microscopy, magnetic tweezers, optical stretcher and optical tweezers pointing out both their advantages and limitations. Finally, we give an outlook on the future of physical probing of cells.

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Active microrheology with optical tweezers: a versatile tool to investigate anisotropies in intermediate filament networks

T Neckernuss, L K Mertens, I Martin, T Paust, M Beil and O Marti

Mechanical properties of cells are determined by the cytoskeleton and especially by intermediate filaments (IFs). To measure the contribution of IFs to the mechanics of the cytoskeleton, we determine the shear moduli of in vitro assembled IF networks consisting of keratin 8/18 and MgCl2, serving as a crosslinker. In this study we want to present a new method, a combination of active and passive microrheology, to characterize these networks. We also show the applicability of the new method and discuss new findings on the organization and force transmission in keratin networks gained by the new method. We trap and move embedded polystyrene particles with an optical tweezers setup in an oscillatory manner. The amplitude response of the trapped particle is measured and evaluated with a lock-in approach in order to suppress random motions. With this technique we determine the degree of isotropy of the assembled network and sense preferred directions due to inhomogeneities of the network. Furthermore, we show that we can deliberately create anisotropic networks by adjusting the assembly process and chamber geometry. To determine whether there are local network anisotropies in a globally isotropic network, we altered the evaluation method and included the motion of embedded particles in the vicinity of the trapped one. The correlations of the observed motions enable us to map local network anisotropies. Finally, we compare mechanical properties determined from passive with ones from active microrheology. We find the networks measured with the active technique to be approximately 20% more compliant than the ones from passive measurements.

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Generation of microfluidic flow using an optically assembled and magnetically driven microrotor

J Köhler, R Ghadiri, S I Ksouri, Q Guo, E L Gurevich and A Ostendorf

The key components in microfluidic systems are micropumps, valves and mixers. Depending on the chosen technology, the realization of these microsystems often requires rotational and translational control of subcomponents. The manufacturing of such active components as well as the driving principle are still challenging tasks. A promising all-optical approach could be the combination of laser direct writing and actuation based on optical forces. However, when higher actuation velocities are required, optical driving might be too slow. Hence, a novel approach based on optical assembling of microfluidic structures and subsequent magnetic actuation is proposed. By applying the optical assembly of microspherical building blocks as the manufacturing method and magnetic actuation, a microrotor was successfully fabricated and tested within a microfluidic channel. The resulting fluid flow was characterized by introducing an optically levitated measuring probe particle. Finally, a freely moving tracer particle visualizes the generated flow. The tracer particle analysis shows average velocities of 0.4–0.5 µm s−1 achieved with the presented technology.

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Inward and outward membrane tubes pulled from giant vesicles

Raktim Dasgupta and Rumiana Dimova

Membrane nanotubes are extruded from giant unilamellar lipid vesicles using a controlled hydrodynamic flow and membrane-attached beads manipulated via optical tweezers. Within a single experiment, the technique can be used to assess various important mechanical and rheological characteristics of the membrane such as the bending rigidity, tension and intermonolayer slip. The application of small flow velocities leads to the extrusion of tubes with sufficiently large diameters conveniently measurable under an optical microscope. For the first time, we show that by suitably controlling the medium flow, inward tubes inside the vesicles can be formed. This approach offers great potential for studying tubulation mechanisms in membrane systems, exhibiting positive as well as negative spontaneous curvatures and should offer a more realistic model for biomembranes because the vesicle membrane tension can adapt freely.

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Optimization of particle trapping and patterning via photovoltaic tweezers: role of light modulation and particle size

J Matarrubia, A Garc?a-Caba?es, J L Plaza, F Agull?-L?pez and M Carrascosa
The role of light modulation m and particle size on the morphology and spatial resolution of nano-particle patterns obtained by photovoltaic tweezers on Fe?:?LiNbO3 has been investigated. The impact of m when using spherical as well as non-spherical (anisotropic) nano-particles deposited on the sample surface has been elucidated. Light modulation is a key parameter determining the particle profile contrast that is optimum for spherical particles and high-m values (m?~?1). The minimum particle periodicities reachable are also investigated obtaining periodic patterns up to 3.5??m. This is a value at least one order of magnitude shorter than those obtained in previous reported experiments. Results are successfully explained and discussed in light of the previous reported models for photorefraction including nonlinear carrier transport and dielectrophoretic trapping. From the results, a number of rules for particle patterning optimization are derived.

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