Freshney Animal Cell Culture Ebook Download
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This eagerly awaited edition reviews the increasing diversity of the applications of cell culture and the proliferation of specialized techniques, and provides an introduction to new subtopics in mini-reviews. New features also include a new chapter on cell line authentication with a review of the major issues and appropriate protocols including DNA profiling and barcoding, as well as some new specialized protocols. Because of the continuing expansion of cell culture, and to keep the bulk of the book to a reasonable size, some specialized protocols are presented as supplementary material online.
R. IAN FRESHNEY, PHD, was an honorary Senior Research Fellow at the Institute of Cancer Sciences at the University of Glasgow, UK. Dr Freshney, who died in 2019, was a world-renowned cancer biologist and a pioneer in cell culture techniques who made important contributions to new approaches for treating cancer patients. He taught cell culture courses at national and international level, and wrote and edited numerous books, including the first seven editions of Culture of Animal Cells. Permissions Request permission to reuse content from this site
The common conception of cytotoxicity is that the cell is killed by the cytotoxin and the assays employed tend to reflect this. There are, however, several distinct aspects of cytotoxicity, differing in cellular mechanisms, outcome, and, consequently, in the assay of their activity (Freshney, 1904). As requirements for in vitro assays become more demanding, driven by mechanistic studies, economics, and by the desire to reduce animal experimentation, there is a need to look more closely at specific cellular responses to toxins and employ an assay most suited to the type of response that is expected. Cytotoxins may have reversible or irreversible effects, and their effects may be immediate or delayed by up to several weeks. There are major differences between (1) physico-chemical damage, which may produce an instantaneous loss of viability, (2) an environmental or pharmaceutical cytotoxin which may have a slight but progressive effect on metabolism over a period of hours or longer, and (3) a loss of reproductive potential, e.g. as a result of irradiation, which may not be immediately apparent in a reduction in the viability of the cells.
Primary cell culture provides more biologically relevant data than that generated using cell lines. Concerns over the use of cell lines have resulted in a growing need for primary cells in a variety of applications from basic research to drug discovery. Often, primary cells are combined with newer technologies such as 3D cell culture given a recent surge within the research community to use better reagents to improve research.
But what are primary cells and how do they differ from cell lines How can you optimize your primary cell culture or create more physiologically relevant models using primary cells in 3D Find the answers to these questions and a comprehensive introduction into the topic here on our primary cell application page, packed with a broad collection of publications, webinars and other helpful resources.
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For majority of the primary cell types, classical medium is not sufficient to support growth, or to retain the phenotypic markers. Our BulletKitTM Media is formulated with additional growth factors and hormones to optimally support consistent growth of primary cells while maintaining the tissue-specific characteristics. Our archive of publications and supporting white papers show suitability of our Primary Cell BulletKitTM Media for establishing complex co-culture models or developing advanced cell culture models.
Whether you need different donors, want to develop a new 3D model, or set up a co-culture, we can help you overcome your primary cell culture challenges. With Lonza's primary cells and media products, you get access to:
Lonza offers a broad collection of cryopreserved primary rat and mouse neuronal cells. Some cell types, for example neural cells, are hard to obtain. Using our primary neuronal cells acquired from mouse and rat brains avoids the prohibitive burden and restrictions regarding animal availability and dissection. A typical challenge with isolating neural cells is to ensure cells have been acquired from specific regions of the brain. Lonza does extensive quality control tests to ensure the neural cells acquired are tissue-specific for the regions of brains specified.
Cell culture refers to the removal of cells from an animal or plant and subsequent cultivation in an artificial environment for scientific research. The first cell culture techniques were developed over 100 years ago and since then have contributed to tremendous breakthroughs in science. Today, it is a fundamental tool used in laboratories around the world for studying the normal physiology and biochemistry of cells, mechanisms underlying disease, including cancer, and effects of drugs and toxic compounds. It is also used in drug screening and development and large-scale manufacturing of biological compounds, such as vaccines and therapeutic proteins.1,2 There's also a growing role of cell culture in the food industry both for testing for contaminants and in cellular agriculture and cultured meat production to ease environmental burdens.
To conduct research requiring cell culture work and to perform fundamental cell culture protocols, there are several key pieces of equipment and some basic reagents that are required, summarized in Tables 1 and 2.
Additional equipment includes an aspirator pump in the laminar flow hood to remove media and reagents from cell culture vessels easily; an autoclave to sterilize equipment and reusable glassware; syringes, needles and forceps; timers; several 70% ethanol spray bottles and paper towel rolls for sterilizing surfaces and equipment; tape and permanent markers for labeling; tube racks and waste bins. 4,5
There are several important design considerations required for any cell culture setup. The most important aspect is using design to maintain an aseptic and sterile environment to prevent the contamination of cells. Firstly, a separate enclosed room or laboratory with one entry/exit point should be used. Hand wash sinks with soap and sanitizer should be close by for hand cleaning on entry and exit of the lab. Dedicated cell culture lab coats and safety goggles should be stored at the lab entrance. The laminar flow hood and incubators should be away from the entrance to minimize contamination risk. It is also important to position the hood and incubator away from any air conditioning units to prevent potentially contaminated airflow from entering the sterile work environment and incubators. There should be ample clear work surfaces that need to be sterilized regularly with sufficient storage space to ensure the surfaces remain clear. All necessary equipment and consumables should be accessible within the laboratory to prevent exiting and re-entering. An ergonomic environment is important for the laminar flow hood with sufficient room for drawers or moveable trolleys of consumables to be at hand when working as well as easy access to the incubator, microscope and centrifuge.6 It's also important to use appropriate plastic consumables that minimize the risk of extractable compounds leaching out and contaminating your cell cultures.
A cell culture laboratory poses risks associated with handling and manipulating cells and tissues as well as toxic, corrosive or mutagenic solvents and reagents. Therefore, adherence to standard microbiological practices and techniques is of paramount importance to mitigate risks and ensure safety at all times. There are four ascending levels of biosafety containment, referred to as biosafety levels (BSL). Each level has standard microbiological practices, safety equipment and facility safeguards to be implemented when dealing with hazardous biomaterials and agents. BSL-1 is the basic level of protection common to most research and clinical laboratories where the agents used are not known to cause disease in normal, healthy humans. BSL-2 is appropriate for moderate-risk agents known to cause human disease of varying severity by ingestion or through percutaneous or mucous membrane exposure. Most cell culture labs should be at least BSL-2, but the exact requirements depend upon the biomaterials used and the type of work conducted. BSL-3 is required for agents that pose a serious and potentially lethal infection and BSL-4, the highest containment level, is required for laboratories working with infectious agents that pose a high individual risk of life-threatening disease.4,7
The following is a list of basic safety recommendations for a cell culture laboratory. The list is by no means complete and should be supplemented with the appropriate biosafety level recommendations.- Always wear appropriate personal protective equipment (PPE) including laboratory coat, gloves and safety goggles.- Always read the material safety data sheet (MSDS) for any substance you are working with to ensure appropriate safety precautions when handling.- Decontaminate all work surfaces before and after your experiments.- Clean laboratory equipment routinely, even if it is not contaminated.- Avoid the creation of aerosols and/or splashes.- Wash your hands after working with potentially hazardous materials and before leaving the laboratory.- Decontaminate all potentially infectious materials before disposal.- Report any incidents that may result in exposure to infectious materials to appropriate personnel (e.g., laboratory supervisor, safety officer).- Do not eat, drink, smoke, handle contact lenses, apply cosmetics, or store food for human consumption in the laboratory. 153554b96e
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