Fluid control structures in microfluidic devices

Methods and apparatus for implementing microfluidic analysis devices are provided. A monolithic elastomer membrane associated with an integrated pneumatic manifold allows the placement and actuation of a variety of fluid control structures, such as structures for pumping, isolating, mixing, routing, merging, splitting, preparing, and storing volumes of fluid. The fluid control structures can be used to implement a variety of sample introduction, preparation, processing, and storage techniques

The Role of Micro fluidic Systems in Biological and Medical Sciences

Micro fluidics is a young discipline. During its beginning, it was mainly an academic field in where researchers study the behavior of fluids at micro scale and how it can be modified with operative and experimental variables. Then, the focus was placed on studying device fabrication process and how to optimize them to lower costs and time, and to enhance system features. After a period of maturation, micro fluidic researchers began to evaluate system usefulness and the possibility of use them in different areas. Micro fluidics became a multidisciplinary field combining concepts of biological and medical sciences and engineering. Diagnostic test, micro particles fabrications, contaminant detection, and medical analyses were first goals. Then, its uses expanded exponentially to other areas opening a world of possibilities. With the advances in miniaturization and material sciences as well as the boom in micro and nanotechnology, manufacturing process became highly precise. New applications in biochemistry, biotechnology, biology and medical sciences were appearing attracting the interest of the industrial sector. Since then, projects are aimed to develop micro fluidic systems with industrial applications. The present contribution describes the characteristics of the three major type of micro fluidic systems, chip-based, capillary-based and paper-based systems. Advantages and limitations of each one are mentioned. In addition, their most important applications in biological and medical sciences are presented.Fil: Helbling, Ignacio Marcelo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Desarrollo Tecnológico Para la Industria Química. Universidad Nacional del Litoral. Instituto de Desarrollo Tecnológico Para la Industria Química; ArgentinaFil: Luna, Julio Alberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Desarrollo Tecnológico Para la Industria Química. Universidad Nacional del Litoral. Instituto de Desarrollo Tecnológico Para la Industria Química; Argentin

Capillary and microdevice electrophoretic tools for genetic analyses: heteroduplex analysis for tuberculosis drug susceptibility and ligase detection reaction for colorectal cancer detection

The traditional format of electrophoresis, the slab gel, is quickly being replaced by capillary and microdevice platforms, which offer improvements in cost, resolution, speed, quantitation and automation in genetic analyses. These techniques also employ a variety of separation matrices while the slab gel is limited to agarose and polyacrylamide. The research presented here explores the use of these electrophoretic formats for the detection of single nucleotide mutations using two genetic models associated with human disease. Slab gel based heteroduplex analysis (HDA), a popular mutation scanning method, uses a specially designed universal heteroduplex generator (UHG) containing controlled variation to enhance the subtle conformational differences caused by single-base substitutions that are difficult to discriminate. Here, the slab gel based HDA-UHG method has been modified to capillary and microdevice formats for the quantitative and reproducible analysis of single-base substitutions in rpoB that give rise to the rifampin-resistant phenotype of Mycobacterium tuberculosis. The capillary method reduced analysis time from 2.5 hours to approximately 30 minutes. The microdevice further reduced analysis time to 6 minutes while maintaining efficiency and resolution. Both capillary and microdevice methods employed methyl cellulose as the sieving matrix at 0.3% and 0.75% (w/v) concentrations, respectively. In colorectal cancer, base substitutions in the K-ras gene occur early in development, are preserved throughout the course of tumor progression and thus, can be used as biomarkers for the diagnosis of early, curable tumors. The current detection scheme uses slab gel electrophoresis and a mutation specific method, multiplexed polymerase chain reaction/ligase detection reaction, to identify all 19 possible single-base substitution mutations at codons 12, 13 and 61. This technique was also adapted and optimized to the capillary and microdevice formats. This study evaluated capillary methods employing both cross-linked as well as entangled polymer matrices. The capillary methods ranged from 30 to 45 minutes in analysis time. The cross-linked capillary exhibited increased deterioration at longer electrokinetic injection times, while severe injection biases were observed in the entangled polymers evaluated. Initial microdevice experiments were possible in approximately 5 minutes using the entangled polymer matrices and have great potential in microdevice analysis for such mutations

Advances in Low Volume Sample Analysis Using Microfluidic Separation Techniques

During the last decades, a great interest has been shown for miniaturised separation techniques. The use of microfluidic techniques fulfills the constant needs for increasing sample throughput and analysis sensitivity, while reducing costs and sample volume consumption. In this chapter, three microfluidic separation techniques will be addressed: capillary electrophoresis, gas chromatography and liquid chromatography. A special attention will be paid to miniaturised liquid chromatography, with a deep investigation of its advantages compared with classical liquid chromatography. Sample preparation adapted to low volumes (a few µl) will also be discussed

A modular approach to high throughput microsystems

A modular high throughput microsystem was developed using microfabrication technology and nucleic acid analysis. The purpose of developing this microsystem is to identify acute infectious disease and prevent contagious outbreak in a matter of time. This microsystem included three major components, a reagent distribution device, a 96 CFPCR array, and a multi-zone thermal system, to efficiently amplify specific DNA fragments to determine the disease status or precisely pinpoint a disease from multiple patients. Polymerase chain reaction (PCR) is the key component in the nucleic acid analysis for disease because it can be used to amplify interested DNA fragments by repeatedly thermal cycles. Temperature distribution is critical to the PCR reaction, and thermal management was studied; the approaches which reached good biochemical results were applied to the design of a 96 CFPCR array and a multi-zone thermal system. Protein adsorption is another issue when PCR is realized in a micro device because of the increased surface-to-volume ratio, which might terminate the biochemical reaction. A series of experiments were carried out to understand the protein adsorption in the microchannels with different geometries. A disposable 96 CFPCR was designed and fabricated on a polycarbonate substrate by double-sided hot embossing and its requirement of multiple temperature zones was fulfilled by building a multi-zone thermal system. Their thermal performance was characterized by numerical simulations and validated by infrared camera experiments. To evenly distribute the analyte with reducing pipetting steps, a reagent distribution device was designed to assemble with the 96 CFPCR array by using passive alignment structures to perform a systematic performance. The passive alignment structures including three pairs of v-groove and hemisphere-tipped post was studied and modified to ensure a smooth passage for mass, momentum, and energy of chemical analyte. Biochemical experiments demonstrated parallel amplifications of both identical and different DNA fragments from the multiple CFPCRs on the multi-zone thermal system, which implied the potential to detect acute infectious disease with acceleration, accuracy, specificity, and high throughput

Microfluidics in Haemostasis: a Review

Haemostatic disorders are both complex and costly in relation to both their treatment and subsequent management. As leading causes of mortality worldwide, there is an ever-increasing drive to improve the diagnosis and prevention of haemostatic disorders. The field of microfluidic and Lab on a Chip (LOC) technologies is rapidly advancing and the important role of miniaturised diagnostics is becoming more evident in the healthcare system, with particular importance in near patient testing (NPT) and point of care (POC) settings. Microfluidic technologies present innovative solutions to diagnostic and clinical challenges which have the knock-on effect of improving health care and quality of life. In this review, both advanced microfluidic devices (R&D) and commercially available devices for the diagnosis and monitoring of haemostasis-related disorders and antithrombotic therapies, respectively, are discussed. Innovative design specifications, fabrication techniques, and modes of detection in addition to the materials used in developing micro-channels are reviewed in the context of application to the field of haemostasi

Bioanalytical methods for studies of homocysteine and novel cardiovascular disease indicators

This dissertation explores the development of analytical methods for studies of CVD biomarkers and related biomolecular indicators. Initially, spectroscopic studies were conducted to investigate the chemical reactivity of homocysteine (Hcy), an independent CVD risk factor and serological biomarker. Consequently, we proposed an alternate theory for in vivo Hcy clearance based on spontaneous pyridoxal tetrahydrothiazine (PT) formation from Hcy and pyridoxal. The validity of PT-assisted Hcy clearance was further evaluated by use of capillary electrophoretic methods, which allowed rapid monitoring of protein oligomerization in PT-protein reaction mixtures. The results of these studies suggest that PT formation is a plausible mechanism for Hcy clearance. Moreover, PT formation was shown to protect proteins from post-translational modification by homocysteine thiolactone. This dissertation also addresses the need for rapid and direct detection methods for CVD biomarkers. Accordingly, we introduced the first plasmon resonant GNP sensing scheme for protein homocystamide. The nanosensor provides visual conformation of protein homocystamide (N-Hcy-protein) by way of a red-to-blue color change. Further sensor investigations conducted with protein nanobioconjugates revealed that the GNP sensing mechanism is dependent on several complex physiochemical and biomolecular interactions including nanoparticle self-assembly, interparticle disulfide cross-linking, and modification-induced protein conformational changes. This dissertation also continues previous atherosclerotic tissue characterization studies by demonstrating the feasibility of using hybrid organic-immunoaffinity extraction for GC-MS analysis of polycyclic aromatic hydrocarbons in human heart plaque samples. This body of work is significant because it proposes new bioanalytical technologies that could enhance CVD screening and treatment