Nanofluidic Immobilization and Growth Detection of Escherichia coli in a Chip for Antibiotic Susceptibility Testing.

Infections with antimicrobial resistant bacteria are a rising threat for global healthcare as more and more antibiotics lose their effectiveness against bacterial pathogens. To guarantee the long-term effectiveness of broad-spectrum antibiotics, they may only be prescribed when inevitably required. In order to make a reliable assessment of which antibiotics are effective, rapid point-of-care tests are needed. This can be achieved with fast phenotypic microfluidic tests, which can cope with low bacterial concentrations and work label-free. Here, we present a novel optofluidic chip with a cross-flow immobilization principle using a regular array of nanogaps to concentrate bacteria and detect their growth label-free under the influence of antibiotics. The interferometric measuring principle enabled the detection of the growth of Escherichia coli in under 4 h with a sample volume of 187.2 µL and a doubling time of 79 min. In proof-of-concept experiments, we could show that the method can distinguish between bacterial growth and its inhibition by antibiotics. The results indicate that the nanofluidic chip approach provides a very promising concept for future rapid and label-free antimicrobial susceptibility tests.

Aberration-corrected cryoimmersion light microscopy.

Cryogenic fluorescent light microscopy of flash-frozen cells stands out by artifact-free fixation and very little photobleaching of the fluorophores used. To attain the highest level of resolution, aberration-free immersion objectives with accurately matched immersion media are required, but both do not exist for imaging below the glass-transition temperature of water. Here, we resolve this challenge by combining a cryoimmersion medium, HFE-7200, which matches the refractive index of room-temperature water, with a technological concept in which the body of the objective and the front lens are not in thermal equilibrium. We implemented this concept by replacing the metallic front-lens mount of a standard bioimaging water immersion objective with an insulating ceramic mount heated around its perimeter. In this way, the objective metal housing can be maintained at room temperature, while creating a thermally shielded cold microenvironment around the sample and front lens. To demonstrate the range of potential applications, we show that our method can provide superior contrast in Escherichia coli and yeast cells expressing fluorescent proteins and resolve submicrometer structures in multicolor immunolabeled human bone osteosarcoma epithelial (U2OS) cells at [Formula: see text]C.

Continuous high throughput nanofluidic separation through tangential-flow vertical nanoslit arrays.

Nanofluidic devices exhibit unique, tunable transport properties that may lead to breakthroughs in molecular separations and sensing. However, the throughput of these devices is orders of magnitude too small for the processing of macroscopic samples. Here we overcome this problem by combining two technological innovations. First, nanofluidic channels are made as vertical slits connecting the two sides of a silicon nitride membrane. Arbitrary arrays of such nanoslits down to 15 nm wide with <6 Å uniformity were made by merging the idea of templating with chemical mechanical polishing to create a scalable, nanolithography-free wafer level process. Second, we provide for efficient solute transport to and from the openings of the nanoslits by incorporating the nanofluidic membrane into a microfluidic tangential-flow system, which is also fabricated at wafer level. As an exemplary application, we demonstrate charge-based continuous flow separation of small molecules with a selectivity of 100 and constant flux over more than 100 hours of operation. This proves the exciting possibility of exploiting transport phenomena governed by precision-engineered nanofluidic devices at a macroscopic scale.

Palladium nanoparticle deposition via precipitation: a new method to functionalize macroporous silicon.

We present an original two-step method for the deposition via precipitation of Pd nanoparticles into macroporous silicon. The method consists in immersing a macroporous silicon sample in a PdCl2/DMSO solution and then in annealing the sample at a high temperature. The impact of composition and concentration of the solution and annealing time on the nanoparticle characteristics is investigated. This method is compared to electroless plating, which is a standard method for the deposition of Pd nanoparticles. Scanning electron microscopy and computerized image processing are used to evaluate size, shape, surface density and deposition homogeneity of the Pd nanoparticles on the pore walls. Energy-dispersive x-ray spectroscopy (EDX) and x-ray photoelectron spectroscopy (XPS) analyses are used to evaluate the composition of the deposited nanoparticles. In contrast to electroless plating, the proposed method leads to homogeneously distributed Pd nanoparticles along the macropores depth with a surface density that increases proportionally with the PdCl2 concentration. Moreover EDX and XPS analysis showed that the nanoparticles are composed of Pd in its metallic state, while nanoparticles deposited by electroless plating are composed of both metallic Pd and PdO x .

Metal electrode integration on macroporous silicon: pore distribution and morphology.

: In this work, a new approach for the one-step integration of interdigitated electrodes on macroporous silicon substrates is presented. Titanium/gold interdigitated electrodes are used to pattern p-type silicon substrates prior the anodization in an organic electrolyte. The electrolyte characteristics, conductivity, and pH have been found to affect the adherence of the metal layer on the silicon surface during the electrochemical etching. The impact of the metal pattern on size distribution and morphology of the resulting macroporous silicon layer is analyzed. A formation mechanism supported by finite element simulation is proposed.