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Whether the question is one of basic cell survival, or whether it is being used to correlate cell number to some other factor such as matrix synthesis, an estimate of cell viability is universally required. In Mammalian Cell Viability: Methods and Protocols, experts in the field describe methods from the most basic which can be performed in any laboratory, to some which require specific pieces of equipment. Initially focusing on methods for monolayer and suspension cells, later chapters describe methods for determining viability within tissue sections and 3 dimensional culture systems. Finally, methods requiring highly specialized equipment are described in order to explain what is possible. Written in the highly successful Methods in Molecular BiologyTM series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and vital tips on troubleshooting and avoiding known pitfalls. Practical and adaptable, Mammalian Cell Viability: Methods and Protocols serves as a self-contained laboratory manual useful to both experienced researchers and those new to this incredibly important and influential field.
Offers a comprehensive overview of cell culture engineering, providing insight into cell engineering, systems biology approaches and processing technology In Cell Culture Engineering: Recombinant Protein Production, editors Gyun Min Lee and Helene Faustrup Kildegaard assemble top class authors to present expert coverage of topics such as: cell line development for therapeutic protein production; development of a transient gene expression upstream platform; and CHO synthetic biology. They provide readers with everything they need to know about enhancing product and bioprocess attributes using genome-scale models of CHO metabolism; omics data and mammalian systems biotechnology; perfusion culture; and much more. This all-new, up-to-date reference covers all of the important aspects of cell culture engineering, including cell engineering, system biology approaches, and processing technology. It describes the challenges in cell line development and cell engineering, e.g. via gene editing tools like CRISPR/Cas9 and with the aim to engineer glycosylation patterns. Furthermore, it gives an overview about synthetic biology approaches applied to cell culture engineering and elaborates the use of CHO cells as common cell line for protein production. In addition, the book discusses the most important aspects of production processes, including cell culture media, batch, fed-batch, and perfusion processes as well as process analytical technology, quality by design, and scale down models. -Covers key elements of cell culture engineering applied to the production of recombinant proteins for therapeutic use -Focuses on mammalian and animal cells to help highlight synthetic and systems biology approaches to cell culture engineering, exemplified by the widely used CHO cell line -Part of the renowned "Advanced Biotechnology" book series Cell Culture Engineering: Recombinant Protein Production will appeal to biotechnologists, bioengineers, life scientists, chemical engineers, and PhD students in the life sciences.
This work present practical, biotechnological applications of flow cytometry techniques for the study of animal, plant and microbial cells, explaining methodologies for sample preparation, staining and analysis. It discusses cell variability in cell culture processes and shows how the quantitative analysis of heterogeneous populations aids in the biotechnological exploitation of cells.
Modern methods have evolved in tissue engineering to evaluate cell viability (CV) in 3D scaffolds and tissues. These involve either the usage of 3D confocal laser microscopy of live or fixed tissues, or separation of cells from the tissue, either live or fixed, and then their analysis by flow cytometry. Generally, working with live cells has the disadvantage that all the scanning needs to be completed immediately at the end of an experiment. Two different approaches can be distinguished: staining intact cell membranes and staining fixed cells. The entire cytoplasm is stained with amine-reactive dyes (ARDs), these use the principle of dead cell exclusion. Here, we list and compare live-cell versus fixed-cells fluorescence-based methods and also show their limitations, especially when working with autofluorescent or cross-linking materials like silk or genipin-reinforced hydrogels. Microscopic techniques have the advantage over flow cytometry-based methods in that these provide the spatial distribution and morphology of the cells. Calcein AM combined with ethidium-homodimer works for most 3D constructs, where no strong fluorescent background is found on the tissue or scaffold. Frequently, however, concentrations and incubation times need to be adjusted for a specific tissue to ensure diffusion of dyes and optimise emittance for detection.
The editors have enlisted a broad range of experts, including microbial ecologists, physiologists, geneticists, biochemists, molecular biologists, and biochemical engineers, who offer practical experience not found in texts and journals. This comprehensive perspective makes MIMB a valuable "how to" resource, the structure of which resembles the sequence of operation involved in the development of a commercial biological process and product.
​Animal cells are the preferred “cell factories” for the production of complex molecules and antibodies for use as prophylactics, therapeutics or diagnostics. Animal cells are required for the correct post-translational processing (including glycosylation) of biopharmaceutical protein products. They are used for the production of viral vectors for gene therapy. Major targets for this therapy include cancer, HIV, arthritis, cardiovascular and CNS diseases and cystic fibrosis. Animal cells are used as in vitro substrates in pharmacological and toxicological studies. This book is designed to serve as a comprehensive review of animal cell culture, covering the current status of both research and applications. For the student or R&D scientist or new researcher the protocols are central to the performance of cell culture work, yet a broad understanding is essential for translation of laboratory findings into the industrial production. Within the broad scope of the book, each topic is reviewed authoritatively by experts in the field to produce state-of-the-art collection of current research. A major reference volume on cell culture research and how it impacts on production of biopharmaceutical proteins worldwide, the book is essential reading for everyone working in cell culture and is a recommended volume for all biotechnology libraries.
Animal cell technology is a growing discipline of cell biology which aims not only to understand structures, functions and behaviors of differentiated animal cells, but also to ascertain their abilities to be used in industrial and medical purposes. The goal of animal cell technology includes accomplishments of clonal expansion of differentiated cells with useful ability, optimization of their culture conditions, modulation of their ability for production of medically and pharmaceutically important proteins and the application of animal cells to gene therapy, artificial organs and functional foods. This volume gives the readers a complete review of present state-of-the-art in Japan and other countries where this field is well advanced. The Proceedings will be useful for the cell biologists, biochemists, molecular biologists, immunologists, biochemical engineers and other disciplines related to animal cell culture, working in either academic environments or in industries of biotechnology and pharmacy.
Fluorescence methods play a leading role in the investigation of biological objects. They are the only non-destructive methods for investigating living cells and microorganisms in vivo. Using intrinsic and artificial fluorescence methods provides deep insight into mechanisms underlying physiological and biochemical processes. This book covers a wide range of modern methods involved in experimental biology. It illustrates the use of fluorescence microscopy and spectroscopy, confocal laser scanning microscopy, flow cytometry, delayed fluorescence, pulse-amplitude-modulation fluorometry, and fluorescent dye staining protocols. This book provides an overview of practical and theoretical aspects of fluorescence methods and their successful application in the investigation of static and dynamic processes in living cells and microorganisms.