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We present a method for label‐free imaging and sorting of cancer cells in blood, which is based on a dielectrophoretic microfluidic chip and label‐free interferometric phase microscopy. The chip used for imaging has been embedded with dielectrophoretic electrodes, and therefore it can be used to sort the cells based on the decisions obtained during the cell flow by the label‐free quantitative imaging method. Hence, we obtained a real‐time, automatic, label‐free imaging flow cytometry with the ability to sort the cells during flow. To validate our model, we combined into the label‐free imaging interferometer a fluorescence imaging channel that indicated the correctness of the label‐free sorting. We have achieved above 98% classification success and 69% sorting accuracy at flow rates of 4 to 7 mL hr−1. In the future, this method is expected to help in label‐free sorting of circulating tumor cells in blood following an initial state‐of‐the‐art cell enrichment.

Polyelectrolyte multilayers assembled from hyaluronic acid (HA) and poly‐l‐lysine (PLL) are most widely studied showing excellent reservoir characteristics to host molecules of diverse nature; however, thick (HA/PLL)n films are often found cell repellent. By a systematic study of the adhesion and proliferation of various cells as a function of bilayer number ""n"" a correlation with the mechanical and chemical properties of films is developed. The following cell lines have been studied: mouse 3T3 and L929 fibroblasts, human foreskin primary fibroblasts VH‐Fib, human embryonic kidney HEK‐293, human bone cell line U‐2‐OS, Chinese hamster ovary CHO‐K and mouse embryonic stem cells. All cells adhere and spread well in a narrow ""cell‐friendly"" window identify in the range of n = 12-15. At n < 12, the film is inhomogeneous and at n > 15, the film is cell repellent for all cell lines. Cellular adhesion correlates with the mechanical properties of the films showing that softer films at higher ""n"" number exhibiting a significant decrease of the Young's modulus below 100 kPa are weakly adherent to cells. This trend cannot be reversed even by coating a strong cell‐adhesive protein fibronectin onto the film. This indicates that mechanical cues plays a major role for cell behavior, also in respect to biochemical ones.

Wound repair is a quiescent mechanism to restore barriers in multicellular organisms upon injury. In chronic wounds, however, this program prematurely stalls. It is known that patterns of extracellular signals within the wound fluid are crucial to healing. Extracellular pH (pHe) is precisely regulated and potentially important in signaling within wounds due to its diverse cellular effects. Additionally, sufficient oxygenation is a prerequisite for cell proliferation and protein synthesis during tissue repair. It was, however, impossible to study these parameters in vivo due to the lack of imaging tools. Here, we present luminescent biocompatible sensor foils for dual imaging of pHe and oxygenation in vivo. To visualize pHe and oxygen, we used time-domain dual lifetime referencing (tdDLR) and luminescence lifetime imaging (LLI), respectively. With these dual sensors, we discovered centripetally increasing pHe-gradients on human chronic wound surfaces. In a therapeutic approach, we identify pHe-gradients as pivotal governors of cell proliferation and migration, and show that these pHe-gradients disrupt epidermal barrier repair, thus wound closure. Parallel oxygen imaging also revealed marked hypoxia, albeit with no correlating oxygen partial pressure (pO2)-gradient. This highlights the distinct role of pHe-gradients in perturbed healing. We also found that pHe-gradients on chronic wounds of humans are predominantly generated via centrifugally increasing pHe-regulatory Na+/H+-exchanger-1 (NHE1)-expression. We show that the modification of pHe on chronic wound surfaces poses a promising strategy to improve healing. The study has broad implications for cell science where spatial pHe-variations play key roles, e.g. in tumor growth. Furthermore, the novel dual sensors presented herein can be used to visualize pHe and oxygenation in various biomedical fields.

Polyelectrolyte multilayer films have been widely studied over the past decade for bioapplications due to their proven reservoir properties for proteins and peptides, growth factors, nucleic acids, drugs, etc. Thick (micrometer-sized) and soft multilayer films made of biopolymers possess high reservoir capacity but are usually cell repellent. Recently, a new approach has been developed based on physical modification of the films made from hyaluronic acid (HA) and poly-L-lysine (PLL) by coating film surface with gold nanoparticles (AuNP). This allows stiffening of the soft HA/PLL film, but still keeps the main part of the film intact to ensure its reservoir ability. Here we have studied cellular adhesion and permeability of the AuNP-coated films. Different cell types (neuronal SH-SY5Y, L929 fibroblasts, human embryonic kidney HEK 293 cells, and cervical cancer HELA cells) have been examined. Film free regions formed by mechanical scratching have been used as controls for cellular adhesion. For the permeability study small dye carboxyfluorescein (CF) and large macromolecule (PLL) have been utilized. As found in this study, (HA/PLL)(24) films are cell repellent for all considered cell lines. But an improved adhesion to the AuNP-coated films has been observed for all the cells lines except of HEK. AuNP coating leads to formation of a kind of semipermeable layer on the film surface, which allows small molecules such as CF to diffuse fast into the film, while diffusion of large molecules (PLL) is hindered. Being cell adhesive and selectively permeable AuNP-coated multilayer films could serve as a novel platform for film-mediated controlled drug delivery.

The prospect of novel therapeutic approaches has renewed the current interest in the fusion of rare cells, like stem cells or primary immune cells. While conventional techniques are only capable of mass fusion, lab-on-a-chip systems often still lack an acceptable method for making the cells available after processing. Here, we present a microfluidic approach for electrofusion on the single-cell level that offers high control over the cells both before and after fusion. For cell pairing and fusion, we employed dielectrophoresis and AC voltage pulses, respectively. Each cell has been characterized and selected before they were paired, fused and released from the fluidic system for subsequent analysis and cultivation. The successful experimental evaluation of our system was further corroborated by numerical simulations. We obtained fusion efficiencies of more than 30% for individual pairs of mouse myeloma and B cell blasts and showed the proliferating ability of the hybrid cells 3 d after fusion. Since aggregates of more than two cells can be fused, the technique could also be developed further for generating giant cells for low-noise electrophysiology in the context of semi-automated pharmaceutical screening procedures.

Formulation of therapeutic proteins into particulate forms is a main strategy for site-specific and prolonged protein delivery as well as for protection against degradation. Precise control over protein particle size, dispersity, purity, as well as mild preparation conditions and minimal processing steps are highly desirable. It is, however, hard to fit all these criteria with conventional preparation techniques. Here a one-step hard-templating synthesis of microparticles composed of functional, non-denatured protein is reported. The method is based on filling porous CaCO 3 microtemplates with the protein near to its isoelectric point (pI) followed by pH- or EDTA-mediated dissolution of the tempplates. In principle, a wide variety of proteins can be converted into microparticles using this approach. The main requirement is an overlap of the protein insolubility and a template solubility for a certain parameter (here pH or EDTA). Here the formulation of insulin particles is studied in detail and it is shown that particles consisting of high molecular weight protein (catalase) can also be prepared. In this context, the synthesis of CaCO 3 templates with controlled size, the mechanism of the protein microparticle formation and mechanical properties of the microparticles are discussed. For the first time, the fabrication of mesoporous monodispersed CaCO 3 microtemplates with identical porocity but tuned diameter from 3 to 20 m is demonstrated. The protein particle diameter can be adjusted by choosing the appropriate template size that is critical for successful pulmonary delivery of insulin. As a first step towards insulin delivery, the in vitro release of insulin at physiological conditions is studied.

Thermoresponsive polymer-coated surfaces based on poly(2-(2-methoxyethoxy)ethyl methacrylate-co-oliog(ethylene glycol) methacrylate) [P(MEO(2)MA-co-OEGMA)] allow switching between cell attachment and detachment. Here, we investigate the temperature-dependent surface interactions between the polymer coating and a colloidal probe in an aqueous medium by means of atomic force microscopy (AFM) force-distance measurements. The analysis of the adhesion forces from AFM retraction curves indentifies two kinds of regimes for the copolymer at temperatures below and above the lower critical solution temperature (LCST). Whereas at 25 degrees C the surface interactions with the polymer in the swollen state are dominated by repulsive forces, at 37 degrees C the surface interactions switch to attractive forces and a stronger adhesion is detected by AFM. Running several healing/cooling cycles repeatedly shows that switching the surface properties provides reproducible adhesion force values. Time-dependent measurements give insight into the switching kinetics, demonstrating that the cell response is coupled to the polymer kinetics but probably limited by the cellular rearrangements.

The control over cell-surface interactions is a highly important issue for biomedical and biotechnology applications. Novel thermoresponsive PEG (Polyethylene glycol) surface coatings allow convenient control over the cell adhesion and do not interfere with membrane proteins. Here we benchmark the ability of the surfaces to control cell adhesion and quantify the interfacial interactions via colloidal probe atomic force microscopy (AFM).

Most of the plywood used in Germany is imported but, on the other hand, resources of large-diameter timber are available. The potential for new plywood plants was investigated regarding the supply potential with spruce and beech timber from forests in North and South Germany and new processing technologies like e.g. defect clipping and jointing of the veneer ribbon. During peeling tests in two German plywood mills with 170 peel rolls from selected locations complete veneer ribbon surfaces were scanned and digitized. Defects (knots, splits etc.) were detected and statistically evaluated. A software was developed which is capable to simulate several processing methods: E.g. clipping between defects and jointing of clear areas vs. clipping of strips with a fixed width and post-sorting of the strips according to their individual defect frequency. Preliminary results show that simulated yield statistics reflect the visual grade of the logs. Moreover, processing parameters like defect tolerance, minimum clear area between defects and grade specifications can be changed to predict and optimize the yield for each peel roll.

At the Keck Smart Materials Integration Laboratory at Penn State University, low-temperature co-fired ceramic (LTCC) material systems have been used to fabricate a number of devices for a variety of applications. This article presents an overview of the integration of the concepts and materials that we have used to achieve miniaturization and unique device function. Examples of microwave filters, metamaterial antennas, and a dielectrophoretic cell sorter will be presented, with emphasis on device modeling and design, prototype construction methods, and test results.