3D correlative single-cell imaging utilizing fluorescence and refractive index tomography

Cells alter the path of light, a fact that leads to well-known aberrations in single cell or tissue imaging. Optical diffraction tomography (ODT) measures the biophysical property that causes these aberrations, the refractive index (RI). ODT is complementary to fluorescence imaging and does not require any markers. The present study introduces RI and fluorescence tomography with optofluidic rotation (RAFTOR) of suspended cells, quantifying the intracellular RI distribution and colocalizing it with fluorescence in 3D. The technique is validated with cell phantoms and used to confirm a lower nuclear RI for HL60 cells. Furthermore, the nuclear inversion of adult mouse photoreceptor cells is observed in the RI distribution. The applications shown confirm predictions of previous studies and illustrate the potential of RAFTOR to improve our understanding of cells and tissues.Comment: 15 pages, 5 figure

Investigating the temperature dependence of the viscosity of a non-Newtonian fluid within lithographically defined microchannels.

We present a study of the rheological phenomenology of a non-Newtonian glass former within hybrid microchannels above the vitrification region. We determined the temperature behavior of the viscosity, which is well fitted by a Vogel-Fulcher-Tamman law for shear rates between 4×10−2 and 9×10−1s−1. The microflow viscosity was compared with previously reported conductivity data of the investigated molecular system. Our findings provide an insight into the coupling between the structural dynamics in the bulk and that within the microchannels, suggesting lithographically defined microfluidic systems as promising tools for the investigation of the rheological properties of complex liquids

Accurate evaluation of size and refractive index for spherical objects in quantitative phase imaging

Measuring the average refractive index (RI) of spherical objects, such as suspended cells, in quantitative phase imaging (QPI) requires a decoupling of RI and size from the QPI data. This has been commonly achieved by determining the object's radius with geometrical approaches, neglecting light-scattering. Here, we present a novel QPI fitting algorithm that reliably uncouples the RI using Mie theory and a semi-analytical, corrected Rytov approach. We assess the range of validity of this algorithm in silico and experimentally investigate various objects (oil and protein droplets, microgel beads, cells) and noise conditions. In addition, we provide important practical cues for future studies in cell biology.Comment: 14 pages, 10 figures, 1 tabl

Ultraviolet-based bonding for perfluoropolyether low aspect-ratio microchannels and hybrid devices.

Producing solvent-resistant microfluidic devices is a challenge for analytical chemistry and biochemistry. We demonstrate a simple and low-cost fabrication approach for the realization of solvent-resistant microchannels based on perfluoropolyether elastomers, exhibiting very low aspect ratios (0.01). The strength of the microchannels sealing is evaluated through the maximum internal pressure (1.52 MPa) prior to device failure, due to delamination at the bonded interface. This approach allows the elastic properties of silicone elastomers, suitable for high quality external connections, to be combined with the non-swelling character of perfluoropolyethers

Interplay between shape and roughness in early-stage microcapillary imbibition.

Flows in microcapillaries and associated imbibition phenomena play a major role across a wide spectrum of practical applications, from oil recovery to inkjet printing and from absorption in porous materials and water transport in trees to biofluidic phenomena in biomedical devices. Early investigations of spontaneous imbibition in capillaries led to the observation of a universal scaling behavior, known as the Lucas-Washburn (LW) law. The LW allows abstraction of many real-life effects, such as the inertia of the fluid, irregularities in the wall geometry, and the finite density of the vacuum phase (gas or vapor) within the channel. Such simplifying assumptions set a constraint on the design of modern microfluidic devices, operating at ever-decreasing space and time scales, where the aforementioned simplifications go under serious question. Here, through a combined use of leading-edge experimental and simulation techniques, we unravel a novel interplay between global shape and nanoscopic roughness. This interplay significantly affects the early-stage energy budget, controlling front propagation in corrugated microchannels. We find that such a budget is governed by a two-scale phenomenon: The global geometry sets the conditions for small-scale structures to develop and propagate ahead of the main front. These small-scale structures probe the fine-scale details of the wall geometry (nanocorrugations), and the additional friction they experience slows the entire front. We speculate that such a two-scale mechanism may provide a fairly general scenario to account for extra dissipative phenomena occurring in capillary flows with nanocorrugated walls

Photocontrolled wettability changes in polymer microchannels doped with photochromic molecules

The authors demonstrate the possibility to control the fluid flow inside microfluidic networks by photoresponsive capillaries. The approach relies on the use of photochromic molecules undergoing reversible changes in their polarity when irradiated with light of specific wavelength, thus varying the wettability of cyclic olefin copolymer microchannels. The realized photoresponsive elements exhibit a decrease up to 20° between the water contact angles of the native and the irradiated surfaces, which could be exploited for enhancing the penetration flow rate of fluids inside microfluidic channels up to 25%. The photocontrollable microfluidic circuitry presents on-off valve behavior, allowing or blocking liquid filling processes on the base of optical control, thus allowing one to manipulate liquid flow within microfluidic networks without mechanical actuation parts

Soft Nanopatterning on Light‐Emitting Inorganic‐Organic Composites

In this work we demonstrate the nanopatterning of nanocomposites made by luminescent zinc oxide nanoparticles and light-emitting conjugated polymers by means of soft molding lithography. Vertical nanofluidics is exploited to overcome the polymer transport difficulties intrinsic in materials incorporating nanocrystals, and the rheology, fluorescence, absolute quantum yield, and emission directionality of the nanostructured composites are investigated. We study the effect of patterned gratings on the directionality of light emitted from the nanocomposites, finding evidence of the enhancement of forward emitted light, due to the printed wavelength-scale periodicity. These results open new possibilities for the realization of nanopatterned devices based on hybrid organic-inorganic systems

Colloidal crystals of compliant microgel beads to study cell migration and mechanosensitivity in 3D

Tissues are defined not only by their biochemical composition, but also by their distinct mechanical properties. It is now widely accepted that cells sense their mechanical environment and respond to it. However, studying the effects of mechanics in in vitro 3D environments is challenging since current 3D hydrogel assays convolve mechanics with gel porosity and adhesion. Here, we present novel colloidal crystals as modular 3D scaffolds where these parameters are principally decoupled by using monodisperse, protein-coated PAAm microgel beads as building blocks, so that variable stiffness regions can be achieved within one 3D colloidal crystal. Characterization of the colloidal crystal and oxygen diffusion simulations suggested the suitability of the scaffold to support cell survival and growth. This was confirmed by live-cell imaging and fibroblast culture over a period of four days. Moreover, we demonstrate unambiguous durotactic fibroblast migration and mechanosensitive neurite outgrowth of dorsal root ganglion neurons in 3D. This modular approach of assembling 3D scaffolds from mechanically and biochemically well-defined building blocks allows the spatial patterning of stiffness decoupled from porosity and adhesion sites in principle and provides a platform to investigate mechanosensitivity in 3D environments approximating tissues in vitro

Nanoparticle image velocimetry at topologically structured surfaces

Nanoparticle image velocimetry ͑nano-PIV͒, based on total internal reflection fluorescent microscopy, is very useful to investigate fluid flows within ϳ100 nm from a surface; but so far it has only been applied to flow over smooth surfaces. Here we show that it can also be applied to flow over a topologically structured surface, provided that the surface structures can be carefully configured not to disrupt the evanescent-wave illumination. We apply nano-PIV to quantify the flow velocity distribution over a polydimethylsiloxane surface, with a periodic gratinglike structure ͑with 215 nm height and 2 m period͒ fabricated using our customized multilevel lithography method. The measured tracer displacement data are in good agreement with the computed theoretical values. These results demonstrate new possibilities to study the interactions between fluid flow and topologically structured surfaces