Download or read book Microfluidic Technologies for Quantitative Single Cell Analysis written by Richard Novak and published by . This book was released on 2013 with total page 251 pages. Available in PDF, EPUB and Kindle. Book excerpt: Multicellular organisms, from roundworms to humans, are composed of interacting individual cells that give rise to an ensemble behavior. Most current technologies enable observation of only the population-level average and often ignore the vast degree of cell heterogeneity present even in clonal populations. Single cell resolution assays of DNA, RNA, proteins, and other biomolecules can yield insights into the complex interactions present in tissues, organs, and whole organisms. Microfluidic systems facilitate single cell analyses by leveraging the micron-scale geometry for improving sensitivity, decreasing reaction time, decreasing reagents consumption, and improving parallelization and automation for high throughput. Microfluidically-generated droplets, in particular, offer extremely high scalability of reactions and straightforward single cell manipulation. This thesis presents the development of microfluidic droplet generator designs and their application for single cell analysis. Developments in microfabricated chip design presented here have resulted in versatile droplet generation tools for a wide range of applications, while novel microfabrication techniques dramatically reduced fabrication time in commercially-viable materials. A radial micropump design increased throughput per device up to 3x106 droplets per hour, allowing us to detect via digital PCR a single pathogenic E. coli O175 in a background of 105 nonpathogenic bacteria. Developing a rapid nickel mold fabrication method has facilitated prototyping and testing of microfluidic designs in thermoplastic materials in as little as 1-3 hours. These microfabrication innovations have accelerated the pace of device development to meet the needs of single cell analysis and other applications. Microfluidic technologies are opening up research paths that so far have been difficult to pursue using conventional methods. High-throughput droplet generation is used to screen purified DNA from healthy subjects exposed to carcinogens for the lymphoma-related t(14;18) chromosomal translocation with a limit of quantitation of less than 1 mutation in 107 genomes and a dynamic range of 105. We also identify unique breakpoint sites and demonstrate the ability to quantify the relative and absolute mutation frequencies within individuals for subjects with multiple mutation events. For analysis of single cell genomes, we present a novel approach for robust DNA purification and analysis using microfluidic agarose droplet encapsulation of single cells. Agarose provides a rigid yet porous shell around cells that enables purification of whole genomes for thousands of cells in parallel without the loss of single cell fidelity. We apply this method to detecting cells containing the t(14;18) translocation and sequencing two DNA targets per cell. This is extended to 9-plex forensic profiling of single cells, thus enabling analysis of complex crime scene samples with multiple contributors or samples with excessive DNA contamination. Finally, droplets are applied to investigating multiple biological parameters per cell, including growth rate, gene expression, and alternative splicing. We perform cell culture in nanoliter droplets for fast generation and monitoring of colonies originating from single cells. Colonies are subsequently assayed for telomerase hTR RNA and hTERT mRNA expression levels and hTERT splice variants. We observe a large degree of expression level bimodality for several splice variants and significant reductions in bimodality coupled with increases in alpha splicing following exposure to sub-lethal concentrations of the anti-cancer compound curcumin. Prospects for microfluidic droplets are discussed in the context of multiparameter single cell analysis as well as applications of single cell analysis to microfluidic organs-on-a-chip. Understanding basic molecular biology mechanisms from the perspective of single cells will yield insights into behavior of multicellular populations with far-reaching scientific and clinical impacts.
Download or read book Microfluidic Technologies for Single Cell Analysis written by Dino Di Carlo and published by . This book was released on 2006 with total page 238 pages. Available in PDF, EPUB and Kindle. Book excerpt: Analysis of single cells is important for a complete understanding of the heterogeneity that is present in cell behavior and function. Current single cell analysis methods provide high throughput information about labeled compounds within cells, but in most cases cannot follow the dynamic processes occurring in signaling pathways. New microfluidic methods have also been developed to assay fast timescale responses or create uniform environments to study cell behavior in a more quantitative manner.
Download or read book Microfluidics for Single Cell Analysis written by Jin-Ming Lin and published by Springer Nature. This book was released on 2019-08-28 with total page 261 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book summarizes the various microfluidic-based approaches for single-cell capture, isolation, manipulation, culture and observation, lysis, and analysis. Single-cell analysis reveals the heterogeneities in morphology, functions, composition, and genetic performance of seemingly identical cells, and advances in single-cell analysis can overcome the difficulties arising due to cell heterogeneity in the diagnostics for a targeted model of disease. This book provides a detailed review of the state-of-the-art techniques presenting the pros and cons of each of these methods. It also offers lessons learned and tips from front-line investigators to help researchers overcome bottlenecks in their own studies. Highlighting a number of techniques, such as microfluidic droplet techniques, combined microfluidics-mass-spectrometry systems, and nanochannel sampling, it describes in detail a new microfluidic chip-based live single-cell extractor (LSCE) developed in the editor’s laboratory, which opens up new avenues to use open microfluidics in single-cell extraction, single-cell mass spectrometric analysis, single-cell adhesion analysis and subcellular operations. Serving as both an elementary introduction and advanced guidebook, this book interests and inspires scholars and students who are currently studying or wish to study microfluidics-based cell analysis methods.
Download or read book Optimization of Trustworthy Biomolecular Quantitative Analysis Using Cyber Physical Microfluidic Platforms written by Mohamed Ibrahim and published by CRC Press. This book was released on 2020-05-31 with total page 335 pages. Available in PDF, EPUB and Kindle. Book excerpt: A microfluidic biochip is an engineered fluidic device that controls the flow of analytes, thereby enabling a variety of useful applications. According to recent studies, the fields that are best set to benefit from the microfluidics technology, also known as lab-on-chip technology, include forensic identification, clinical chemistry, point-of-care (PoC) diagnostics, and drug discovery. The growth in such fields has significantly amplified the impact of microfluidics technology, whose market value is forecast to grow from $4 billion in 2017 to $13.2 billion by 2023. The rapid evolution of lab-on-chip technologies opens up opportunities for new biological or chemical science areas that can be directly facilitated by sensor-based microfluidics control. For example, the digital microfluidics-based ePlex system from GenMarkDx enables automated disease diagnosis and can bring syndromic testing near patients everywhere. However, as the applications of molecular biology grow, the adoption of microfluidics in many applications has not grown at the same pace, despite the concerted effort of microfluidic systems engineers. Recent studies suggest that state-of-the-art design techniques for microfluidics have two major drawbacks that need to be addressed appropriately: (1) current lab-on-chip systems were only optimized as auxiliary components and are only suitable for sample-limited analyses; therefore, their capabilities may not cope with the requirements of contemporary molecular biology applications; (2) the integrity of these automated lab-on-chip systems and their biochemical operations are still an open question since no protection schemes were developed against adversarial contamination or result-manipulation attacks. Optimization of Trustworthy Biomolecular Quantitative Analysis Using Cyber-Physical Microfluidic Platforms provides solutions to these challenges by introducing a new design flow based on the realistic modeling of contemporary molecular biology protocols. It also presents a microfluidic security flow that provides a high-level of confidence in the integrity of such protocols. In summary, this book creates a new research field as it bridges the technical skills gap between microfluidic systems and molecular biology protocols but it is viewed from the perspective of an electronic/systems engineer.
Download or read book Microfluidics 2018 written by Microfluidics 2018, New Technologies and Applications in Biology, Biochemistry and Single-Cell Analysis and published by . This book was released on 2018 with total page pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book Single Cell Analysis in Biotechnology and Systems Biology written by Fan-Gang Tseng and published by MDPI. This book was released on 2018-10-01 with total page 237 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book is a printed edition of the Special Issue "Single Cell Analysis in Biotechnology and Systems Biology" that was published in IJMS
Download or read book Cell Analysis on Microfluidics written by Jin-Ming Lin and published by Springer. This book was released on 2017-10-25 with total page 435 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book presents a detailed overview of the design, formatting, application, and development of microfluidic chips in the context of cell biology research, enumerating each element involved in microfluidics-based cell analysis, discussing its history, status quo, and future prospects, It also offers an extensive review of the research completed in the past decade, including numerous color figures. The individual chapters are based on the respective authors' studies and experiences, providing tips from the frontline to help researchers overcome bottlenecks in their own work. It highlights a number of cutting-edge techniques, such as 3D cell culture, microfluidic droplet technique, and microfluidic chip-mass spectrometry interfaces, offering a first-hand impression of the latest trends in the field and suggesting new research directions. Serving as both an elementary introduction and advanced guidebook, the book interests and inspires scholars and students who are currently studying microfluidics-based cell analysis methods as well as those who wish to do so.
Download or read book Microfluidic Single cell Technologies for Assaying Lymphocyte Interactions written by Burak Dura and published by . This book was released on 2015 with total page 144 pages. Available in PDF, EPUB and Kindle. Book excerpt: Immune cells do not live in isolation but interact to coordinate their many actions. One of the chief routes they foster communication is through direct physical interactions that enables them to read and interpret signals mediated at membrane interfaces. Despite the critical importance of these direct interactions in determining crucial developmental and functional immunological responses, their dynamic nature together with vast heterogeneity and polyfunctionality of individual immune cells have presented technical challenges for their systematic investigation. In particular, only limited tools are available that can exert control over the individual cells and their microenvironments to be able to precisely define interactions and deeply profile their outcomes at the individual cell level to resolve emerging immune responses within each single-cell. To fill this critical void, this thesis presents the development and implementation of novel microfluidic technologies for single-cell analysis of direct cell-cell interactions in immunology. By combining carefully designed weir-based hydrodynamic traps with a multistep cell loading procedure, the microfluidic devices capture and controllably pair hundreds of cells in parallel. This approach provides requisite control over interactions with one-to-one interacting partners, well-defined and synchronous initiation of interactions, and enduring contacts. It also provides full control over the soluble microenvironment by solution exchange without losing cell registration. Accordingly, these features enable monitoring and assaying lymphocyte interactions longitudinally from the beginning with multiparametric single-cell measurements. These capabilities in turn allow probing into complete immune cell activation window from the very onset for direct correlation analyses within hundreds of individual cells in a single experiment. We apply these new 'microfluidic cell pairing' technologies to quantitative investigation of lymphocyte interactions to elucidate lymphocyte activation dynamics and their relation to diverse functional behaviors at the single-cell level. These studies help resolve qualitatively and quantitatively distinct calcium signaling patterns in single CD8 T cells based on varying T cell receptor affinities which correlate with differential cytokine output. Similar studies with natural killer (NK) cells identify a previously unreported inverse correlation between the strength of early calcium signaling and cytokine production, and further indicate a calcium-dependent mechanism for selective regulation of cytotoxicity and cytokine production in NK cells. Collectively, these findings provide essential insight into the regulation and evolution of immune responses within individual immune cells, and establish the potential of these new microfluidic technologies to address important questions on many aspects of cell-cell interactions across biology in general and in immunology in particular.
Download or read book Development of Microfluidic and Biological Technologies for Quantitative and Dynamic Analysis of Gene Expression in Single Cells written by Nils Schneider and published by . This book was released on 2018* with total page pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book Microfluidics for Single cell Analysis written by and published by . This book was released on 2019 with total page 263 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book summarizes the various microfluidic-based approaches for single-cell capture, isolation, manipulation, culture and observation, lysis, and analysis. Single-cell analysis reveals the heterogeneities in morphology, functions, composition, and genetic performance of seemingly identical cells, and advances in single-cell analysis can overcome the difficulties arising due to cell heterogeneity in the diagnostics for a targeted model of disease. This book provides a detailed review of the state-of-the-art techniques presenting the pros and cons of each of these methods. It also offers lessons learned and tips from front-line investigators to help researchers overcome bottlenecks in their own studies. Highlighting a number of techniques, such as microfluidic droplet techniques, combined microfluidics-mass-spectrometry systems, and nanochannel sampling, it describes in detail a new microfluidic chip-based live single-cell extractor (LSCE) developed in the editor's laboratory, which opens up new avenues to use open microfluidics in single-cell extraction, single-cell mass spectrometric analysis, single-cell adhesion analysis and subcellular operations. Serving as both an elementary introduction and advanced guidebook, this book interests and inspires scholars and students who are currently studying or wish to study microfluidics-based cell analysis methods.
Download or read book Biosensors for Single Cell Analysis written by Jian Chen and published by Academic Press. This book was released on 2021-10-26 with total page 184 pages. Available in PDF, EPUB and Kindle. Book excerpt: Biosensors for Single-Cell Analysis explores a wide range of biosensor technologies and their applications in single-cell characterization and analysis. Sections cover key biophysical and chemical single-cell properties that consider proteomic, metabolic, electrical, mechanical and optical properties. Each chapter features key definitions and case studies, providing detailed guidance for researchers who want to replicate covered solutions in their work. Tutorial sections, evaluations of the current state-of-the-field and future developments are also included. Microfluidic approaches to characterization, such as microfluidic impedance flow cytometry and microfluidic flow cytometry are considered alongside more conventional approaches, such as mass spectroscopy, fluorescent and mass flow cytometry. Additionally, key types of biosensors are covered, including atomic force microscopy, micropipette aspiration, optical tweezers, microfluidic hydrodynamic stretchers, microfluidic constriction channel and microfluidic optical stretchers. - Includes chapters focused on key single-cell properties, such as proteomic, metabolic and mechanical characterization - Features case studies that illustrate the application of biosensors for single-cell analysis - Considers microfluidic approaches for each single-cell property discussed - Explores future directions for single-cell analysis and biosensor technology
Download or read book Quantitative Single cell Analysis System Based on Soft Lithography Technologies written by Adrian Yan Ho Lau and published by . This book was released on 2007 with total page 330 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book Single Cell Assays written by Paul C.H. Li and published by Springer Nature. This book was released on 2023-07-10 with total page 227 pages. Available in PDF, EPUB and Kindle. Book excerpt: This detailed volume explores the use of single-cell assays in research for drug discovery, microfluidics, and more. The book delves into methodologies involving a variety of cell types and diseases, small molecules and biologics, as well as studies of the genome and transciptome. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step and readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, Single-Cell Assays: Microfluidics, Genomics, and Drug Discovery serves to enable researchers to obtain a rapid overview in state-of-the-art microfluidic single-cell assays and an impression of what possibilities these assays offer to drug discovery.
Download or read book Open Space Microfluidics written by Emmanuel Delamarche and published by John Wiley & Sons. This book was released on 2018-01-18 with total page 450 pages. Available in PDF, EPUB and Kindle. Book excerpt: Summarizing the latest trends and the current state of this research field, this up-to-date book discusses in detail techniques to perform localized alterations on surfaces with great flexibility, including microfluidic probes, multifunctional nanopipettes and various surface patterning techniques, such as dip pen nanolithography. These techniques are also put in perspective in terms of applications and how they can be transformative of numerous (bio)chemical processes involving surfaces. The editors are from IBM Zurich, the pioneers and pacesetters in the field at the forefront of research in this new and rapidly expanding area.
Download or read book Microfluidic Technologies for Investigating Dynamics based Regulation in S Cerevisiae written by Richard O'Laughlin and published by . This book was released on 2020 with total page 142 pages. Available in PDF, EPUB and Kindle. Book excerpt: Technological advances in molecular biology over the last several decades have enabled researchers to probe the inner workings of living systems at unprecedented scale and resolution. These new capabilities have resulted in the emergence of the field of quantitative biology, which seeks to utilize mathematical modeling along with techniques from systems biology and synthetic biology in order to ascertain the design principles that underlie the structure and function of biological regulatory networks. In pursuit of this goal, it has become clear that heterogeneity at the single-cell level and the dynamics, or time-dependent behavior, of these networks are critical features of a multitude of biological processes. Much of the foundational work establishing the importance of these themes began from the study of microbes such as the budding yeast Saccharomyces cerevisiae. However, even in unicellular organisms such as S. cerevisiae, significant challenges remain in tracking and analyzing single cells over long periods of time, as well as monitoring gene expression dynamics at scale. Development of these capabilities is critical, not only for understanding the cell biology of S. cerevisiae, but also for being able to apply the mechanistic insights obtained from yeast to evolutionarily conserved pathways in higher eukaryotes. In this thesis, I describe the design and application of microfluidic technologies for S. cerevisiae that address these limitations. In Chapter 1, I detail the utility of microfluidic devices and time-lapse fluorescence microscopy for analyzing single cells and recording biological dynamics. In Chapter 2, I describe the development of novel microfluidic devices that enable long-term isolation and tracking of single yeast cells, culminating in a design that can monitor cells over the course of their entire replicative lifespans. In Chapters 3 and 4, I discuss applications of the microfluidic technologies described in Chapter 2. Chapter 3 concerns the discovery of metabolic cycles in flavin fluorescence at the single-cell level, which can oscillate along with and independently of the cell division cycle, persisting even when cellular respiration is blocked. Chapter 4 describes the uncovering of two divergent paths taken by single yeast cells during replicative aging, in which distinct dynamics of heterochromatin silencing at the rDNA region that modulate cellular lifespan can be detected in each group. In Chapter 5, I apply the design principles validated in the construction of microfluidic devices for tracking single cells in order to develop a high-throughput microfluidic platform for recording gene expression dynamics of thousands of yeast strains simultaneously. I demonstrate a proof-of-principle of this technology by monitoring changes in gene expression across more than 4000 yeast strains in real-time during the diauxic shift. The microfluidic devices developed and described herein underscore the importance of single-cell analysis and dynamics-based regulation by elucidating novel sources of heterogeneity at the single-cell level and demonstrating how metabolic, chromatin silencing and gene expression dynamics contribute to the regulation of complex processes such as cell division, replicative aging and growth in changing environments. Further, these technologies establish a foundation upon which future studies can continue to pursue quantitative biology work in yeast, from the single-cell level to the genome scale.
Download or read book Micro Nanofluidic Devices for Single Cell Analysis written by Fan-Gang Tseng and published by MDPI. This book was released on 2018-10-04 with total page 167 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book is a printed edition of the Special Issue "Micro/Nanofluidic Devices for Single Cell Analysis" that was published in Micromachines
Download or read book Open Space Microfluidics written by Emmanuel Delamarche and published by John Wiley & Sons. This book was released on 2018-04-30 with total page 440 pages. Available in PDF, EPUB and Kindle. Book excerpt: Summarizing the latest trends and the current state of this research field, this up-to-date book discusses in detail techniques to perform localized alterations on surfaces with great flexibility, including microfluidic probes, multifunctional nanopipettes and various surface patterning techniques, such as dip pen nanolithography. These techniques are also put in perspective in terms of applications and how they can be transformative of numerous (bio)chemical processes involving surfaces. The editors are from IBM Zurich, the pioneers and pacesetters in the field at the forefront of research in this new and rapidly expanding area.
