Volume 3, Issue 1

A novel microscopic technique termed laser switching contrast microscopy (LSCM) allows for the imaging of the dynamics of optically switchable proteins in single cell compartments. We present an application for the monitoring of diffusive properties of single molecules of the photo-switchable fluorescent protein Dreiklang (DRK). LSCM in the cell nucleus of Chinese hamster ovary (CHO) cells cytoplasmically expressing DRK unravels quick diffusive equilibration of the DRK molecules inside the whole cytoplasm and inside the cell nucleus within seconds. The nuclear membrane is also highly permeable for DRK. Inside the nucleus entirely distinct regions are found that only partially enable diffusive protein redistribution with mean square displacement proportional to time while in other regions the mobility of the proteins seems to be restricted. After photo-switching string like patterns of light DRK molecules are observed in the cell nucleus. In addition a fraction of these DRK molecules appears immobile. The findings support recent theories of the cell interior described as a random obstacle model with an additional immobile fraction of DRK. Numerical simulations show that at different illumination intensity and different distance from the laser focus similar patterns for fluorescence recovery might be obtained in spite of strongly varying diffusion constants.

Surface plasmon resonance biosensors have recently known a rapid diffusion in the biological field and a large variety of sensor configurations is currently available. Biological applications are increasingly demanding sensor miniaturization, multiple detection in parallel, temperature-controlled environment and high sensitivity. Indeed, versatile and tunable sensing platforms, together with an accurate biological environment monitoring, could improve the realization of custom biosensing devices applicable to different biological reactions. Here we propose a smart and high throughput fabrication protocol for the realization of a custommicrofluidic plasmonic biochip that could be easily tuned and modified to address different biological applications. The sensor chip here presented shows a high sensing capability, monitored by an accurate signal calibration in the presence of concentration and temperature variation.

We present a method for controllably producing longitudinal acoustic trapping sites inside microfluidic channels. Air bubbles are injected into a micro-capillary to create bubble-bounded ‘micro-cavities’. A cavity mode is formed that shows controlled longitudinal acoustic trapping between the two air/water interfaces along with the levitation to the centre of the channel that one would expect from a lower order lateral mode. 7 μm and 10 μm microspheres are trapped at the discrete acoustic trapping sites in these micro-cavities.We show this for several lengths of micro-cavity.

Fluorescence correlation spectroscopy relies on temporal autocorrelation analysis of fluorescence intensity fluctuations that spontaneously arise in systems at equilibrium due to molecular motion and changes of state that cause changes in fluorescence, such as triplet state transition, photoisomerization and other photophysical transformations, to determine the rates of these processes. The stability of a fluorescent molecule against dark state conversion is of particular concern for chromophores intended to be used as reference tags for comparing diffusion processes on multiple time scales. In this work, we analyzed properties of two fluorescent proteins, the photoswitchable Dreiklang and its parental eGFP, in solvents of different viscosity to vary the diffusion time through the observation volume element by several orders of magnitude. In contrast to eGFP, Dreiklang undergoes a dark-state conversion on the time scale of tens to hundreds of microseconds under conditions of intense fluorescence excitation, which results in artificially shortened diffusion times if the diffusional motion through the observation volume is sufficiently slowed down. Such photophysical quenching processes have also been observed in FCS studies on other photoswitchable fluorescent proteins including Citrine, from which Dreiklang was derived by genetic engineering. This property readily explains the discrepancies observed previously between the diffusion times of eGFP- and Dreiklang-labeled plasma membrane protein complexes.

This paper describes the selective infiltration and storage of picoliter volumes of water and IPA in arrays of sealed SU-8 microwells. Microwells, with a volume of approximately 300 picoliters, are fabricated employing photolithography and a polymer onto polymer lamination method to seal the structures with a thin cover of SU-8 and PDMS in order to suppress the evaporation of the infiltrated liquids. A glass capillary is used to punch through the SU-8/PDMS cover and to infiltrate the liquid of interest into the microwells. The influence of the mixing ratio of the PDMS and its curing agent is studied and the results show that a lower ratio of 2:1 suppresses the evaporation more when compared to the standard mixing ratio of 10:1. In regards to water and IPA, the dwell time in the reservoirs was increased by approximately 50 % and 450 % respectively. Depending on the physical properties of the microwells and the liquids, the SU-8/PDMS cover suppresses the evaporation up to 32 mins for water and 463 mins for IPA, respectively, until the microwell is completely empty again. Additionally, multiple infiltrations of the same microwell are demonstrated using two immiscible liquids IPA and paraffin oil. Based on the popular polymers SU-8 and PDMS, the sealed microwell structures are scalable and combinable with different glass capillaries according to the needs of future analytical research and medical diagnostics.

The article reports, in the general context of developing techniques to generate microjets, nanojets and even individual nanodroplets, a new method to obtain such formations by interaction of a single laser pulse at 532 nm with an individual/single mother droplet in pendant position in open air. The beam energy per pulse is varied between 0.25 and 1 mJ, the focus diameter is 90 μm, and the droplet’s volumes are either 3 μl or 3.5 μl. Droplet’s shape evolution and jet emission at impact with laser pulse was visualised with a high speed camera working at 10 kfps. Reproducible jets and/or separated microdroplets and nanodroplets are obtained which shows potential for applications in particular in jet printing. It is demonstrated that it becomes possible to play with the geometrical symmetry of both laser excitation and liquid in order to manage the number and the orientation of an induced microjet and consequently to actuate the orientation and the production of nanodroplets by light.

Most of the vibrating mechanisms of optofluidic systems are based on local heating of membranes that induces liquid flow.We report here a new type of diaphragm pump in a liquid film based on the optical radiation pressure force. We modulate a low power laser that generates, at resonance, a symmetric vibration of a free standing soap film. The film lifetime strongly varies from 56 s at low power (2 mW) to 2 s at higher power (70 mW). Since the laser beam only acts mechanically on the interfaces, such a pump could be easily implemented on delicate microequipment on chips or in biological systems.

We report two viable strategies to assemble and manipulate arrays of nano- and micro-particles by means of topological defects (TDs) in anisotropic fluids. Exploiting different boundary conditions, single TD, 1D arrays of TDs are tailored in liquid crystal twist cells. In a first approach, light-guided control of particles captured in disclination lines is demonstrated involving the use of a photosensitive chiral dopant within a nematic host. In the second one, an applied voltage enables a continuous displacement and deformation of the particles arrays. The reported results open up new possibilities for managing nano- and micro-metric objects over large distances.

The nonlinear optical properties of two graphene derivatives, graphene oxide and graphene fluoride, are investigated by means of the Z-scan technique employing 35 ps and 4 ns, visible (532 nm) laser excitation. Both derivatives were found to exhibit significant third-order nonlinear optical response at both excitation regimes, with the nonlinear absorption being relatively stronger and concealing the presence of nonlinear refraction under ns excitation, while ps excitation reveals the presence of both nonlinear absorption and refraction. Both nonlinear properties are of great interest for several photonics, opto-fluidics, opto-electronics and nanotechnology applications.