Search

The process of hydraulic fracture is well known in both natural (e.g. veining and mineralisation) and engineered environments (e.g. stimulating tight mudrocks and sandstones to boost their hydraulic properties). Here, we report a method and preliminary data that simulates both tensile fracture and fluid flow at elevated pressures. To achieve this we developed a sample assembly consisting of a cylindrical core drilled with an axial borehole encapsulated in a 3D printed jacket permitting fluid from the borehole to move through the freshly generated tensile fracture to a voluometer. The permeability of Nash Point Shale increases from a pre-fracture value of 10 -18 to 10 -20  m 2 (1 microDarcy, μD to 0.01 μD) to 2 × 10 -15  m 2 (2 milliDarcy, mD) immediately after fracture (at 2.1 MPa confining pressure). Permeability is strongly dependent on confining pressure, decreasing to 0.25 × 10 -15  m 2 (0.25 mD) at 19 MPa confining pressure (approximately 800 m depth), and does not recover when confinement is removed. Using concomitant measurements of the radial strain as a proxy for fracture aperture, we conclude that the effective permeability is governed solely by the width of the developed cracks, revealed by post-test X-Ray Computed Tomography to be planar, extending radially from the central conduit.

The transport of bioactive compounds to the site of action is a great challenge. A promising approach to overcome application-related problems is the development of targeting colloidal transport systems, such as micelles which are equipped with uptake mediating moieties. Here, we investigated a set of novel lipopeptides which exhibit a surfactant-like structure due to attachment of two palmitoyl chains to the N-terminus of cationic or anionic amino acid sequences. We analyzed the association behavior of these lipopeptides by using 5(6)-carboxyfluorescein (CF)-labeled derivatives as a fluorescent probe and different spectroscopic methods such as fluorescence anisotropy and fluorescence correlation spectroscopy (FCS). The photophysical properties as well as the diffusion and rotational movements of the CF-labeled lipopeptides were exploited to determine the cmc and the size of the micelles consisting of lipopeptides. We could distinguish cationic and anionic lipopeptides by their association behavior and by studying the interactions with mouse brain capillary endothelial cells (b.end3). The cationic derivatives turned out to be very strong surfactants with a very low cmc in the micromolar range (0.5-14 μM). The unique combination of micelle-forming property and cell-penetrating ability can pave the road for the development of a novel class of efficient drug carrier systems.

Ammonium serves as key nitrogen source and metabolic intermediate, yet excess causes toxicity. Ammonium uptake is mediated by ammonium transporters, whose regulation is poorly understood. While transport can easily be characterized in heterologous systems, measuring transporter activity in vivo remains challenging. Here we developed a simple assay for monitoring activity in vivo by inserting circularly-permutated GFP into conformation-sensitive positions of two plant and one yeast ammonium transceptors ('AmTrac' and 'MepTrac'). Addition of ammonium to yeast cells expressing the sensors triggered concentration-dependent fluorescence intensity (FI) changes that strictly correlated with the activity of the transporter. Fluorescence-based activity sensors present a novel technology for monitoring the interaction of the transporters with their substrates, the activity of transporters and their regulation in vivo, which is particularly valuable in the context of analytes for which no radiotracers exist, as well as for cell-specific and subcellular transport processes that are otherwise difficult to track. DOI:http://dx.doi.org/10.7554/eLife.00800.001.

Time-resolved emission data (fluorescence decay and fluorescence depolarization) of two three-color Förster resonance energy transfer (tc-FRET) systems consisting of a carbostyril donor (D), a ruthenium complex (Ru) as relay dye, and a Cy5 derivative (Cy) or, optionally, an anthraquinone quencher (Q) were carefully analyzed using advanced distribution analysis models. Thereby, it is possible to get information on the flexibility and mobility of the chromophores which are bound to double stranded (ds) DNA. Especially the distance distribution based on the analysis of the fluorescence depolarization is an attractive approach to complement data of fluorescence decay time analysis. The distance distributions extracted from the experimental data were in excellent agreement with those determined from accessible volume (AV) simulations. Moreover, the study showed that for tc-FRET systems the combination of dyes emitting on different time scales (e.g., nanoseconds vs microseconds) is highly beneficial in the distribution analysis of time-resolved luminescence data in cases where macromolecules such as DNA are involved. Here, the short lifetimes can yield information on the rotation of the dye molecule itself and the long lifetime can give insight in the overall dynamics of the macromolecule.

We report on a new three-color FRET system which we were able to verify in peptides as well as in synthetic DNA. All three chromophores could be introduced by a building block approach avoiding postsynthetic labeling. Additional features are robustness, matching spectroscopic properties, high-energy transfer, and sensitivity. The system was investigated in detail on a set of peptides as well as an array of tailored oligonucleotides. The detailed analysis of the experimental data and comparison with theoretical considerations were in excellent agreement. It is shown that in the case of polypeptides specific interaction with the fluorescence probes has to be considered. In contrast with DNA, the fluorescence probes did not show any indications of such interactions. The novel three-color FRET toolbox revealed the potential for applications studying fundamental processes of three interacting molecules in life science applications.