J. Michael Köhler

Chip devices for miniaturized biotechnology

Chip devices were introduced in chemistry and molecular biology to improve the read-out of information from molecular systems by efficient analytical procedures and to organize automated experiments. Biochips and chip reactor systems are of interest for cellular processes, too, and can be regarded as components in interfaces for the information exchange between living nature and digital electronic systems. In this minireview, different types of chip reactors for biotechnological applications like nanotiterplates, chip thermocyclers and devices for segmented flow operations are discussed. Finally, an outlook is given on the application of chip reactor systems, which are promising tools for automated experiments with highly parallelized screening procedures, for artificial microcompartmentation, cell analogue systems, micro-ecological studies, investigations on modulated morphogenesis, and for a bioanalogue molecular nanotechnology.

Metallic nanowires created by biopolymer masking

Nanometer-scale structures are of great interest as potential building blocks for future electronic devices. One significant challenge is the electrical connection of such blocks by nanowires. We present here the use of filamentous biopolymer structures (microtubules) as a template for the creation of metallic nanowires. The microtubules served as a mask in an etching step of a thin metal layer. This approach results in formation of metallic paths (wires) of well-defined dimensions, with a shape determined by the biological template. Electrical measurements confirmed the realization of conductive metallic nanowires.

Polyacrylamid/silver composite particles produced via microfluidic photopolymerization for single particle-based SERS microsensorics.

A micro-continuous-flow process was applied for the preparation of swellable polyacrylamide particles incorporating silver nanoparticles. These sensor particles are formed from a mixture of a colloidal solution of silver nanoparticles and monomer by a droplet-based procedure with in situ photoinitiation of polymerization and a subsequent silver enforcement in batch. The obtained polymer composite particles show a strong SERS effect. Characteristic Raman signals of aqueous solutions of adenine could be detected down to 0.1 μM by the use of single sensor particles. The chosen example demonstrates that the composite particles are suitable for quantitative microanalytical procedures with a high dynamic range (3 orders of magnitude for adenine).

Formation of star-like and core-shell AuAg nanoparticles during two- and three-step preparation in batch and in microfluidic systems

Regular dendrit-like metal nanoparticles and core-shell nanoparticles were formed by the reduction of mixtures of tetrachloroaurate and silver nitrate solutions with ascorbic acid at room temperature in two- and three-step procedures. The formation of these particles was found in batch experiments as well as in micro flow-through processes using static micromixers. The characteristic diameters of 4-branched star particles were in the range between 60 and 100 nm. The typical particles consist of four metal cores which are embedded in a common shell. Additionally, particles with five and more metallic cores were formed, to some extent, and aggregates of the 4-branched particles also were formed. Larger aggregates and network-like structures of connected star particles were formed after sedimentation. The properties of the formed particles are dependent on the educt concentrations as well as on the order of mixing steps and on the time interval between them. Obviously, the relation of nucleation and particle growth in relation to the concentrations of metal ions determines the composition and the properties of formed nanoparticles. So, starlike particles are observed in case of nucleation of Au in absence of silver ions but with silver deposition after short nucleation time. Spherical core shell particles are formed in case of silver salt addition after complete reduction of tetrachloroaurate in flow-through experiments with sufficient residence time between both mixing steps. Polymer layers are always found in the form of a second outer shell even if the polymer solutions are added in an early stage of particle formation.

Single-Step Microfluidic Synthesis of Various Nonspherical Polymer Nanoparticles via in Situ Assembling: Dominating Role of Polyelectrolytes Molecules

In this paper, a microfluidic approach has been used for the synthesis of ellipsoidal, dumbbell, rodlike, and necklacelike polymer nanoparticles. High yields of special types of nonspherical nanoparticles have been achieved by the implementation of an emulsion polymerization into microfluidic arrangement with a micro hole-plate reactor for the formation of monomer droplets. Here, in particular, the formation of nonspherical polymer nanoparticles is dependent on the presence of polyelectrolyte surface active molecules such as poly(4-styrenesulfonic acid-co-maleic acid) sodium salt (PSS-co-PM), poly(sodium-p-styrenesulfonate) (PSSS), and polyanetholesulfonic acid sodium salt (PAES). The shapes and sizes of the interparticle nanoassemblies are precisely controlled by adjusting the concentration of polyelectrolytes in the aqueous phase, and by choosing suitable flow rate ratios (aqueous to monomer phase), respectively. The formation of polymer nanoparticles with different morphologies can be explained by a spontaneous in situ assembling under partial electrostatic repulsive control in the single step synthesis. The effect of particle charge and the competition between thermal motion of particles and electrostatic repulsion on the spontaneous assembling under the condition of a limited polarizability are discussed here as an important factor for the formation process of nonspherical polymer nanoparticles.

Multiscale materials from microcontinuous-flow synthesis: ZnO and Au nanoparticle-filled uniform and homogeneous polymer microbeads

Tri(propylene glycol) diacrylate (TPGDA) was found to be an excellent monomer for the stabilization and dispersion of inorganic nanoparticles. Uniform nano-Au/poly(TPGDA) and nano-ZnO/poly(TPGDA) composite microbeads were synthesized in situ using a designed axisymmetric capillary-based flow-focusing microfluidic device without any additional surfactant or coupling agent. Using the designed mixing-enhanced microfluidic device, homogeneous nano-inorganic/polymer composites with a high content of nanoparticles were obtained. Morphologies of the composites were characterized by SEM, TEM, surface microscopy, dark-field microscopy and internal fluorescence.

Making electrical contact to single molecules

We are interested in the characterization of electron transport phenomenon through single (bio)molecules. A prerequisite for electrical measurements is the contacting of molecules in the nanometer range. This paper describes various steps toward this goal, starting from statistically distributed molecules (namely, microtubules) adsorbed on a microstructured surface. Monitored by scanning force microscopy and video-enhanced microscopy, adsorption parameters were studied and optimized, including protein-mediated adsorption, induced orientation, and controlled desorption. Line structures connecting the adsorbed molecules to prestructured electrodes were created by means of electron beam-induced deposition (EBD) in a scanning electron microscope. It is shown how such EBD lines can be used as masks for the structuring of metal contacts, and an electrical characterization of these contacts is given.

Microfluidic Assisted Synthesis of Multipurpose Polymer Nanoassembly Particles for Fluorescence, LSPR, and SERS Activities

Potential biomedical applications such as controlled delivery with sustained drug release profile demand for multifunctional polymeric particles of precise chemical composition and with welldefined physicochemical properties. The real challenge is to obtain the reproducible and homogeneous nanoparticles in a minimum number of preparation steps. Here, single-step nanoarchitectures of soft surface layered copolymer nanoparticles with a regular tuning in the size via micro flow-through assisted synthesis are reported. Interfacial copolymerization induces the controlled compartmentalization where a hydrophobic core adopts spherical shape in order to minimize the surface energy and simultaneously shelter in the hydrophilic shelllike surface layer. Surface layer can swell in the aqueous medium and allow controlled entrapping of functional hydrophobic nanoparticles in the hydrophilic interior via electrostatic interaction which can be particularly interesting for combined fluorescence activity. Furthermore, the nanoarchitecture of size and concentration controlled polymer-metal nanoassembly particles can be implemented as an ideal surface-enhanced Raman scattering substrate for detection of the trace amounts of various analytes.

Cultivation of Chlorella vulgaris in microfluid segments and microtoxicological determination of their sensitivity against CuCl2 in the nanoliter range

The cultivation of the monocellular green alga Chlorella vulgaris was implemented into microfluid segments to demonstrate the possibility of an automated screening of toxic effects of the common algaecide CuCl2. Therefore, the nutritional as well as light and carbon dioxide requirements of the algae had to be adapted to the microfluidic device. Generally, sequences of about 350 fluid segments with single volumes of about 500 nL were applied for the dose–response experiments. The growth of algae cultures inside microfluidic segments was non‐invasively measured by microflow through techniques using two different optical channels. A multi‐endpoint detection was realized by the photometric characterization of cell density by transmission measurements and the measurement of density of autofluorescent cells. The different methods revealed comparable half maximal effective concentrations (EC50) in the range between 34.6 and 39.9 μg/mL for the toxicity of CuCl2 to the green algae C. vulgaris. By reference experiments in microtiter plates lower EC50 were achieved presumably caused by increased alkalinity of the growth medium due to higher photosynthesis. The results show that the microsegmented flow technique is well suited for the automated determination of dose/response functions for microorganisms like C. vulgaris and for the application of multi‐endpoint procedures at the nanoliter scale.

Control of shape and size of polymer nanoparticles aggregates in a single-step microcontinuous flow process: a case of flower and spherical shapes.

Controlled aggregation of polymer nanoparticles for building anisotropic nano- and microstructures via a self-assembling bottom-up process is an important strategy. Therefore, in this work, the formation of structured poly(methyl methacrylate) (PMMA) particles with diameters between lower micrometer and submicrometer range by use of a microcontinuous flow arrangement was investigated in the presence of nonionic water-soluble polymer polyvinylpyrrolidone (PVP). The investigations show that the microreaction strategy is well applicable and allows a tuning of size and shape of nanoparticles in dependence on reactant concentrations and flow rate ratios. Larger and complex structured polymer particles have been found at lower PVP concentration, whereas more compact submicron-sized particles were formed at higher PVP concentrations. The addition of ionic surfactants modulates the generation of characteristic particle shapes. The observation of intermediate states between complex flowerlike particles and simple spheres in dependence on the applied concentration of low molecular weight surfactants supports the explanation of particle formation by a mechanism with superposition of particle growth and assembling. When mixed surfactants (PVP-SDS or PVP-CTAB) are used, the final particles shape depends on the concentration of individual concentrations of surfactants and on the competition between mobility, solvation, and micelle formations.