Tommaso F. Bersano-Begey

Integrated elastomeric components for autonomous regulation of sequential and oscillatory flow switching in microfluidic devices

A critical need for enhancing usability and capabilities of microfluidic technologies is the development of standardized, scalable, and versatile control systems1,2. Electronically controlled valves and pumps typically used for dynamic flow regulation, although useful, can limit convenience, scalability, and robustness3–5. This shortcoming has motivated development of device-embedded non-electrical flow-control systems. Existing approaches to regulate operation timing on-chip, however, still require external signals such as timed generation of fluid flow, bubbles, liquid plugs or droplets, or an alteration of chemical compositions or temperature6–16. Here, we describe a strategy to provide device-embedded flow switching and clocking functions. Physical gaps and cavities interconnected by holes are fabricated into a three-layer elastomer structure to form networks of fluidic gates that can spontaneously generate cascading and oscillatory flow output using only a constant flow of Newtonian fluids as the device input. The resulting microfluidic substrate architecture is simple, scalable, and should be applicable to various materials. This flow-powered fluidic gating scheme brings the autonomous signal processing ability of microelectronic circuits to microfluidics where there is the added diversity in current information of having distinct chemical or particulate species and richness in current operation of having chemical reactions and physical interactions.

Individually programmable cell stretching microwell arrays actuated by a Braille display

Cell culture systems are often static and are therefore nonphysiological. In vivo, many cells are exposed to dynamic surroundings that stimulate cellular responses in a process known as mechanotransduction. To recreate this environment, stretchable cell culture substrate systems have been developed, however, these systems are limited by being macroscopic and low throughput. We have developed a device consisting of 24 miniature cell stretching chambers with flexible bottom membranes that are deformed using the computer-controlled, piezoelectrically actuated pins of a Braille display. We have also developed efficient image capture and analysis protocols to quantify morphological responses of the cells to applied strain. Human dermal microvascular endothelial cells (HDMECs) were found to show increasing degrees of alignment and elongation perpendicular to the radial strain in response to cyclic stretch at increasing frequencies of 0.2, 1, and 5 Hz, after 2, 4, and 12h. Mouse myogenic C2C12 cells were also found to align in response to the stretch, while A549 human lung adenocarcinoma epithelial cells did not respond to stretch.

Microfluidic platform for chemotaxis in gradients formed by CXCL12 source-sink cells

Chemokine CXCL12 promotes CXCR4-dependent chemotaxis of cancer cells to characteristic organs and tissues, leading to metastatic disease. This study was designed to investigate how cells expressing CXCR7 regulate chemotaxis of a separate population of CXCR4 cells under physiologic conditions in which cells are exposed to gradients of CXCL12. We recapitulated a cancer-stroma microenvironment by patterning CXCR4-expressing cancer cells in microchannels at spatially defined positions relative to CXCL12-producing cells and CXCR7-expressing cells. CXCR7 scavenges and degrades CXCL12, which has been proposed to facilitate CXCR4-dependent chemotaxis through a source-sink model. Using the microchannel device, we demonstrated that chemotaxis of CXCR4 cells depended critically on the presence and location of CXCR7 cells (sink) relative to chemokine secreting cells (source). Furthermore, inhibiting CXCR4 on migrating cells or CXCR7 on sink cells blocked CXCR4-dependent chemotaxis toward CXCL12, showing that the device can identify new therapeutic agents that block migration by targeting chemoattractant scavenging receptors. Our system enables efficient chemotaxis under much shallower yet more physiological chemoattractant gradients by generating an in vitro microenvironment where combinations of cellular products may be secreted along with formation of a chemoattractant gradient. In addition to elucidating mechanisms of CXCL-12 mediated chemotaxis, this simple and robust method can be broadly useful for engineering multiple microenvironments to investigate intercellular communication.

Micro- and nanotechnologies for studying cellular function.

The study of complex biological systems requires methods to perturb the system in complex yet controlled ways to elucidate mechanisms and dynamic interactions, and to recreate in vivo conditions in flexible in vitro set-ups. This paper reviews recent advances in the use of micro- and nanotechnologies in the study of complex biological systems and the advantages they provide in these two areas. Particularly useful for controlling the chemical and mechanical microenvironments of cells is a set of techniques called soft lithography, whereby elastomeric materials are used to transfer and generate micro- and nanoscale patterns. Examples of some of the capabilities of soft lithography include the use of elastomeric stamps to generate micropatterns of protein and the use of elastomeric channels to localize chemicals with subcellular spatial resolutions. These types of biological micro- and nanotechnologies combined with mathematical modeling will propel our understandings of cellular and subcellular physiology to new heights.

Uniform cell seeding and generation of overlapping gradient profiles in a multiplexed microchamber device with normally-closed valves

Generation of stable soluble-factor gradients in microfluidic devices enables studies of various cellular events such as chemotaxis and differentiation. However, many gradient devices directly expose cells to constant fluid flow and that can induce undesired responses from cells due to shear stress and/or wash out of cell-secreted molecules. Although there have been devices with flow-free gradients, they typically generate only a single condition and/or have a decaying gradient profile that does not accommodate long-term experiments. Here we describe a microdevice that generates several chemical gradient conditions on a single platform in flow-free microchambers which facilitates steady-state gradient profiles. The device contains embedded normally-closed valves that enable fast and uniform seeding of cells to all microchambers simultaneously. A network of microchannels distributes desired solutions from easy-access open reservoirs to a single output port, enabling a simple setup for inducing flow in the device. Embedded porous filters, sandwiched between the microchannel networks and cell microchambers, enable diffusion of biomolecules but inhibit any bulk flow over the cells.

Evolving feature-extraction algorithms: adapting genetic programming for image analysis in geoscience and remote sensing

Discusses a relatively new procedure in the computer-assisted design of pattern-extraction algorithms. The procedure involves the adaptation of genetic programming, a recent technique that has been used for automatic programming, for image processing and analysis. This paper summarizes several of the measures the authors have taken to develop two prototype systems that help a user to design pattern-extraction algorithms.

Musica ex Machina: Composing 16th-Century Counterpoint with Genetic Programming and Symbiosis

GPmuse is software which explores one connection between computation and creativity using a symbiosis-inspired genetic programming paradigm in which distinct agents collaborate to produce 16th-century counterpoint.

Ice roughness classification and ERS SAR imagery of Arctic sea ice: evaluation of feature-extraction algorithms by genetic programming

This paper describes a validation of accuracy associated with a recent algorithm that has been designed to extract ridge and rubble features from multiyear ice. Results show that the algorithm performs well with low-resolution ERS SAR data products.

A Java collaborative interface for genetic programming applications: image analysis for scientific inquiry

Discusses several key issues involved in designing and using a Java collaborative interface for genetic programming applications over the World Wide Web. We present our implementation that has been used in a new system that assists scientists in classifying and extracting novel features in remotely sensed satellite imagery. This paper also identifies issues in developing a class library that facilitates rapid prototyping of such collaborative graphical user interfaces for genetic programming, and suggests how other researchers could benefit from them.

Label-free biosensing using a photonic crystal structure in a total-internal-reflection geometry.

A novel optical biosensor using a one-dimensional photonic crystal structure in a total-internal-reflection geometry (PCTIR) is presented and investigated for label-free biosensing applications. This simple configuration forms a micro Fabry- Perot resonator in the top layer which provides a narrow optical resonance to enable label-free, highly sensitive measurements for the presence of analytes on the sensing surface or the refractive index change of the surrounding medium in the enhanced evanescent field; and at the same time it employs an open sensing surface for real-time biomolecular binding detection. The high sensitivity of the sensor was experimentally demonstrated by bulk solvent refractive index changes, ultrathin molecular films adsorbed on the sensing surface, and real-time analytes binding, measuring both the spectral shift of the photonic crystal resonance and the change of the intensity ratio in a differential reflectance measurement. Detection limits of 7×10-8 RIU for bulk solvent refractive index, 6×10-5 nm for molecular layer thickness and 24 fg/mm2 for mass density were obtained, which represent a significant improvement relative to state-ofthe- art surface-plasmon-resonance (SPR)-based systems. The PC-TIR sensor is thus seen to be a promising technology platform for high sensitivity and accurate biomolecular detection.