Micro and Nanosystems - Volume 2, Issue 4, 2010

For decades, the scientists have been trying to solve the intricate questions of the biology and the human science by reaching successful results. Although many questions are still open and one of the main reasons is the inadequate methods that biologists and biochemists have available. To date a snapshot of the actual scientific world shows a huge gap between biology and technology and a strong difficulty in talking, due to the different background and language. However, even if slowly, the scenario is progressively changing and part of the scientific community, albeit small, is promoting the interdisciplinary interaction between biological and technological world. To understand biology, where still complexity is unrevealed, it is urgent to understand and to aware that novel and intricate technologies are needed. In fact, even if it is clear that biology requires that even more complex and exciting questions are solved, sometimes the biologists are unfamiliar with what could be out there to help answer, whilst, usually, such technologies are developed by people who have not in depth knowledge of biological problems. The efforts of those developing key technologies are not optimal when they are not supported by an interdisciplinary knowledge of the biological problem space. Micro total analysis systems seem to be one of the most useful tools for supporting the biological research and several commercial systems are based on microfluidic principles to perform routinely analysis. There are several reasons why microfluidic systems are so important for biology. First, as widely shown, the size of such systems allows scaling down of the process parameters with clear advantages in terms of the characteristic time and efficiency of the analysis. Secondly, high precision control of small volumes of sample makes these systems particularly useful when it is necessary to interrogate biological samples. Finally, microfluidic systems allow the microenvironment where biological samples, especially cells, normally live to be mimicked. Microfluidic systems allow the micro- and nano-topology of the systems to be finally tailored. To be effective, care needs to be taken that the developed microfluidic system designed to mimic the in vivo conditions does not modify the phenotype and genotype of the cells. The potential for microfluidic science to answer key biological questions is the most exciting development since its first applications 20 years ago in analytical chemistry. The most authoritative examples of the success include (i) the potential to grow tissue from human cells inside the microfluidic systems with stimulation able to mimic the in vivo conditions, (ii) isolation and screening of circulating tumor cells with a dual advantage of diagnosis and prognosis, and (iii) interrogation of cells for the screening of small chemical molecules. My own view is that most of the important open questions of life need to be solved by models and systems that allow simultaneously to simplify the complexity of the in vivo studies and to keep invariant the surrounding conditions. Microfluidic systems can be an aid to develop such models. The content of this issue exemplifies some of what it is intended to achieve in terms of synergic collaboration between biology and technology. • Microfabrication of microfluidic systems- Specific application realized for responding to the needs of cells and for cell culturing in mimetic bioenviroment. • Standard Operations- Identification of tumor cells behavior and interaction in controlled biochemical environment. • Application- A patch clamp based chip for neuronal cell interrogation.

We propose a polymer-based microfluidic motherboard that integrates waveguides and fluidic networks providing an interface between microfluidic systems and the outer world. The motherboard facilitates interconnections of several microfluidic chips for multiplexed and simultaneous analysis. It offers a modular network for microfluidic chips, allowing complex microfluidic processes, where each microchip has a particular function. The motherboard includes microfluidic channels machined by micromilling technology and bonded thermally. Waveguides were integrated in the motherboard in the same fabrication process. Additionally, pins were fabricated, which are part of the fluidic interconnections and allow the alignment of a chip with the waveguides of the motherboard. To establish high density microfluidic interconnections between the motherboard and external tubes, reversible PDMS sockets have been utilized. For interconnecting the polymer waveguides to the light source and to the detection system, PDMS optical plugs have been developed. To demonstrate the performance of the device, test structures were designed that contained fluidic channels, waveguides and, additionally, custom made o-rings allowing alignment to the connection pins of the motherboard. The single chips were attached to the motherboard by just pressing them onto the alignment pins. A sealed fluidic connection between chip and motherboard is ensured by the deliberate mismatch in diameter between the o-rings of the chip and the pins of the motherboard. The maximally applicable hydraulic pressure before leakage, as well as propagation and coupling losses for the waveguides and optical interconnections have been characterized and tested.

Spatial localization and handling of cells and colloid trajectories in a microfluidic channel are required to fully control their interactions at a single level. This paper presents a novel transparent multilevel aligned electrode microfluidic chip that is fast to process and reusable, with which high throughput for dielectrophoretic handling of polarizable particles can be achieved. Performances of the chips are evaluated by electrical impedance measurements and light diffusion characterizations. The observation of the Indium Tin Oxyde (ITO) electrode degradation process allows a better understanding of the behavior of ITO at low frequencies. Electrical and optical characterizations confirmed that this process is a redox reaction of indium that seriously alters the conductivity and the transparency of the ITO layer in the microfluidic chip. A safe range of AC-voltage frequencies is given for dielectrophoretic handling. Finally, we demonstrate within this chip focalization and stop-and-go functions for 1 μm polystyrene colloidal particles.

In this article a mathematical model is presented in which the fluid dynamic interaction between the liquid flow in a scaffold and growing cells is simulated. The model is based on a computational fluid dynamic (CFD) model for the representation of the fluid dynamic conditions in the scaffold. It includes furthermore a simple biological growth model based on Michaelis Menten type kinetics for the growth of cells. The model includes biomass, substrate and oxygen as the most important growth limiting components in the system. Furthermore the growth, decay and maintenance respiration of the cells are considered in the model. In a variation of the model the growth of the biomass is influenced by the fluid dynamic induced shear stress level, which the cells are exposed to. In parallel an experimental growth of stem cells has been performed in a 3D perfusion reactor system and the culturing has been stopped after 2, 8 and 13 days. The development of the cells is compared to the simulated growth of cells and it is attempted to draw a conclusion about the impact of the shear stress on the cell growth.

Calcium has been demonstrated to have a fundamental role in cell-cell recognition mediating the carbohydrate- carbohydrate and the protein-carbohydrate interactions. A self assembled multifunctional microfluidic array has been used to exploit the interactions between breast tumor cells and an engineered surface. Engineering concerns the topology of the surface, since microbeads have been used to increase the micro roughness of the surface, the biochemistry, since the microbeads are conjugated by carbohydrates before being integrated inside the microfluidic chamber. Microfluidic environment permits to control parameters such as the shear force and the velocity and mimicking the microenvironment surrounding the cells. To this end it is extremely important to investigate the role of calcium in cell carbohydrate interactions in a microfluidic environment. Investigation has been performed varying the calcium concentration of cell media and the fluidic condition. Breast tumor cells have been used. Static and dynamic experiments have been performed. Static experiments have shown that differences in the cell adhesion behavior are only detectable when dealing with calcium free buffer, whilst the results of adhesion in correspondence of different carbohydrates have been lost when dealing with calcium enriched media. Adhesion of glucose is independent on the composition of the buffer. At 3000 μl sec-1 60 and 80% of adherent cells have been counted in according to the free and enriched calcium media. The trend of the results from the dynamic experiments is not affected by the presence of calcium, and three different conditions can be identified. In particular, between 10 and 15 μl min-1 they adhere and start roll on the substrate at high flow rate. We assume that during the first seconds calcium is not able to drive the interaction between carbohydrates and cells and hydrodynamic phenomena, the balance between shear force and velocity, play the most important role.

Colony formation is one of the most important characteristics of the pluripotency of embryonic stem cells (ESC). On a flat surface of soft material such as polydimethylsiloxane (PDMS), the formation efficiency as well as the processing stability of ESC colonies is significantly lower than that on harder material surfaces. To still use PDMS as substrate for cell culture which is interesting for both microdevice fabrication and cell differentiation control, we deposited single nanofibers on PDMS for enhanced handling of ESCs. As expected, not only much more ESC colonies could be formed and stay on the PDMS surface with nanofibers but also they showed a narrow size distribution as compared to the results obtained on the PDMS surface without nanofibers.

We have previously described the designs of two planar patch-clamp neurochips and their application to the electrophysiological study of molluscan neurons cultured on-chip. Neuron attachment and growth over apertures on the neurochip surface permitted the acquisition of whole-cell patch-clamp recordings. To broaden the application of these neurochips from molluscan to mammalian neurons, we conducted a study of cell-to-aperture interaction to optimize conditions for these smaller, more fragile cells. For this purpose, we designed a “sieve” chip having multiple apertures on its surface. Random growth of rat cortical neurons resulted in a 32% (n = 324) probability of cell growth over 2 μm diameter apertures; larger diameters resulted in growth through the aperture. Based on these findings, single-aperture neurochips were fabricated having 2 μm diameter aperture and preliminary electrophysiological recordings from cortical cultures at 14 DIV are presented. The implications of this study for the next-generation neurochips are discussed.

Focusing on the development of a nanometer scale carrier in the drug-delivery-system for cancer therapy, we prepared a lipid nano-emulsion (LNE) from a lipid mixture of soybean oil (SO), phosphatidylcholine (PC) and sodium palmitate as a vehicle for chloroaluminum phthalocyanine (ClAlPC), a photosensitizer used for the photodynamic treatment of cancer. To elucidate its distribution in our LNE formulation, we proposed a modified three-phase (SO core, PC monolayer and water phase) model using the molar partition coefficients (Kps) of ClAlPC in LNE/water and PC small unilamellar vesicle (PC SUV)/water systems. In the presence of LNE or PC SUV, a spectrophotometric absorption maximum due to monomeric ClAlPC was observed, but was absent in buffered solution because of its property of selfaggregation in aqueous media, showing that the partitioned ClAlPC exists as a monomer. The Kp values of ClAlPC were calculated using the intensity of the absorption maximum in the second derivative spectra. Although the Kp values showed ClAlPC concentration dependence due to the association of ClAlPC in the water phase, they were on the order of 104 for the concentrations of 1-5 μM ClAlPC, indicating that our LNE having a lipid concentration of 72 mM can encapsulate 95-99% of the spiked ClAlPC. Furthermore, most of ClAlPC was found to be located in the PC monolayer surrounding the SO droplets in the LNE. These results indicate that the PC monolayer not only acts as a water-oil interface, but also stores monomeric ClAlPC in our LNE effectively.

With the advancement in ultra precision technologies and micromachining processes, the fabrication of well defined nanochannels has become feasible. Nanochannels have applications in various fields such as biomedical analysis, fuel cell, and water technologies. Understanding of characteristics of fluid transport in nanoscale is currently under research, and new insights have been gained. However, there still are a number of phenomen that have not yet been described by a complete theory. Experimental investigations provide interesting results which are unique in the nanoscale and require extensive studies to be understood. This paper aims to provide a general view on the transport phenomena in nanochannels from the different aspects of theory, experiment, fabrication and simulation. The major results of this field in the recent years are also highlighted. This review contributes to the preliminary understanding of the emerging research fields of nanofluidics.

Recent advances in microfluidics and microfabrication techniques have led to a variety of portable and inexpensive lab-on-a-chip devices to make quantitative assays of microscale and nanoscale bioparticles. Among them, electrical impedance flow cytometers have become an indispensable tool in clinical and research laboratories for analysis of micro/ nano bio-objects. Because of their simplicity and capability of single cell analysis, electrical impedance flow cytometers have been used widely to detect and characterize latex beads, pollen, biological cells, bacteria, viruses and DNA. One long standing drawback of the electrical impedance flow cytometers is their low throughput, namely they can only process a small amount of analyte at one time. To enable rapid analysis and real time detection of micro and nano objects, high throughput microfluidic electrical impedance flow cytometers have been developed to analyze a large volume of sample in a reasonable time. These devices are especially useful for rapid detection of bio-objects present in ultra low concentrations without a need for tedious preconcentration. In this article, we will review the state-of-the-art high throughput microfluidic electric impedance sensors for rapid analysis of micro bioparticles, including 1) multi-channel Coulter counters, 2) multiplexed resistive pulse sensors, 3) electrical impedance spectroscopy sensors and 4) radio frequency high bandwidth particle counters. Advantages and limitations of each type of impedance sensors are discussed.