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Miles Turner

Animal Cell Culture Book Pdf: A Comprehensive and Practical Guide for Beginners and Experts


Animal Cell Culture Book Pdf




Animal cell culture is a technique that involves growing animal cells in an artificial environment that mimics their natural conditions. Animal cell culture is widely used in biotechnology and medicine for various purposes, such as producing recombinant proteins, antibodies, viral vectors, tissues, organs, and drugs. Animal cell culture also provides a valuable tool for studying the structure, function, and interaction of animal cells, as well as their response to different stimuli and stressors.




Animal Cell Culture Book Pdf



However, animal cell culture is not a simple or straightforward process. It requires careful preparation, maintenance, monitoring, and control of the cells and their environment. It also faces many challenges and limitations, such as contamination, variability, instability, scalability, cost, safety, and ethics. Therefore, it is essential for anyone who wants to work with animal cells in culture to have a solid understanding of the basic principles and practices of animal cell culture.


In this article, we will provide an overview of animal cell culture, covering its techniques, equipment, applications, and resources. We will also introduce some useful books that can help you learn more about animal cell culture in depth. By reading this article, you will gain a comprehensive knowledge of animal cell culture and its importance in biotechnology and medicine.


Animal Cell Culture Techniques




One of the most important aspects of animal cell culture is how to prepare and maintain the cells in a suitable environment that supports their growth, survival, differentiation, and function. This involves selecting the appropriate type of cells, media, supplements, subculturing methods, cryopreservation methods, thawing methods, and contamination prevention and detection techniques.


How to prepare and maintain animal cells in culture?




The first step in preparing animal cells in culture is to obtain them from a reliable source. There are two main types of animal cells that can be used for culturing: primary cells and immortalized cell lines. Primary cells are isolated directly from animal tissues or organs, and they retain most of their original characteristics and functions. However, they have a limited lifespan and can only be cultured for a few passages before they senesce or die. Immortalized cell lines are derived from primary cells that have been transformed or modified to overcome the senescence barrier and proliferate indefinitely. However, they may lose some of their original features and functions, and they may acquire genetic and phenotypic changes over time.


Once the cells are obtained, they need to be cultured in a suitable medium that provides them with the necessary nutrients, growth factors, hormones, and other factors that regulate their metabolism and physiology. There are many types of media available for animal cell culture, such as serum-based media, serum-free media, protein-free media, chemically defined media, and custom-made media. The choice of media depends on the type of cells, the purpose of the culture, and the cost and availability of the components. Generally, serum-based media are the most commonly used media for animal cell culture, as they contain a complex mixture of proteins, lipids, carbohydrates, vitamins, minerals, hormones, and growth factors that support the growth and function of most animal cells. However, serum-based media also have some disadvantages, such as variability, contamination, immunogenicity, and interference with downstream applications. Therefore, some researchers prefer to use serum-free media or protein-free media that contain only the essential components for animal cell culture, or chemically defined media that contain only pure and known components for animal cell culture. These types of media offer more consistency, control, safety, and specificity for animal cell culture, but they may also require more optimization and validation for different cell types and applications.


In addition to the medium, animal cells in culture may also require some supplements to enhance their growth, survival, differentiation, and function. Some common supplements used for animal cell culture are antibiotics, antifungals, antimycotics, amino acids, glutamine, pyruvate, sodium bicarbonate, buffering agents, antioxidants, and attachment factors. The choice and concentration of supplements depend on the type of cells, the type of medium, and the purpose of the culture.


Another important aspect of maintaining animal cells in culture is how to subculture them periodically to prevent overcrowding, nutrient depletion, waste accumulation, and senescence. Subculturing involves transferring a portion of the cells from one culture vessel to another with fresh medium. The frequency and method of subculturing depend on the type of cells, the type of medium, the type of culture vessel, and the growth rate and density of the cells. Generally, there are two main methods for subculturing animal cells: trypsinization and mechanical dissociation. Trypsinization involves treating the cells with an enzyme called trypsin that breaks down the proteins that attach the cells to each other and to the culture surface. This allows the cells to detach easily and form a single-cell suspension that can be transferred to a new culture vessel. Mechanical dissociation involves physically disrupting the cell-cell and cell-surface interactions by using a pipette, a scraper, a needle, or a sieve. This also results in a single-cell suspension that can be transferred to a new culture vessel.


How to prevent and detect contamination in animal cell culture?




One of the biggest challenges and threats in animal cell culture is contamination by microorganisms, such as bacteria, fungi, yeasts, mycoplasmas, viruses, or other animal cells. Contamination can compromise the quality, integrity, and validity of the animal cell culture and its applications. It can also cause serious health risks for the researchers and the end-users of the products derived from animal cell culture. Therefore, it is essential to prevent and detect contamination in animal cell culture by following good laboratory practices and using appropriate techniques and tools.


The best way to prevent contamination in animal cell culture is to use sterile techniques and materials when handling and manipulating the cells and their environment. This includes wearing personal protective equipment such as gloves, gowns, masks, and goggles; using sterile instruments such as pipettes, syringes, needles, scissors, and forceps; using sterile reagents such as media, supplements, water, and ethanol; using sterile containers such as flasks, dishes, tubes, and bottles; using a biosafety cabinet for opening and closing the containers and performing any procedures that may generate aerosols or droplets; using an incubator with a HEPA filter and a UV light to maintain a sterile atmosphere for culturing the cells; and disposing of any waste materials in a biohazard bag or autoclave.


Another way to prevent contamination in animal cell culture is to detect it as soon as possible by using various techniques and tools. Some common signs of contamination in animal cell culture are changes in the appearance, behavior, growth rate, or viability of the cells; changes in the color, clarity, pH, or viscosity of the medium; and presence of visible particles, turbidity, or gas bubbles in the culture. Some common techniques and tools for detecting contamination in animal cell culture are microscopy, staining, culturing, PCR, ELISA, and DNA fingerprinting. These techniques and tools can help identify the type, source, and extent of contamination in animal cell culture and guide the appropriate actions to take to eliminate or minimize the impact of contamination.


Animal Cell Culture Equipment




Another important aspect of animal cell culture is how to use the appropriate equipment and instruments for culturing, handling, observing, and analyzing the cells and their environment. This involves selecting and operating a biosafety cabinet, an incubator, a microscope, a centrifuge, and a bioreactor for animal cell culture. It also involves monitoring and controlling the physical and chemical parameters of animal cell culture, such as temperature, pH, dissolved oxygen, carbon dioxide, osmolality, and glucose.


How to use a biosafety cabinet, an incubator, a microscope, and a centrifuge for animal cell culture?




A biosafety cabinet (BSC) is a ventilated enclosure that provides a sterile and safe work area for performing animal cell culture procedures. A BSC protects the cells from contamination by filtering the incoming and outgoing air through high-efficiency particulate air (HEPA) filters that remove 99.97% of particles larger than 0.3 microns. A BSC also protects the operator and the environment from exposure to potentially hazardous agents by creating a negative pressure inside the cabinet that prevents the escape of aerosols or droplets. A BSC should be located in a quiet and low-traffic area of the laboratory away from doors, windows, vents, fans, or other sources of air turbulence. A BSC should be certified annually by a qualified technician to ensure its proper functioning and safety. A BSC should be cleaned and disinfected before and after each use with 70% ethanol or another suitable disinfectant. A BSC should be operated according to the manufacturer's instructions and following the standard operating procedures (SOPs) of the laboratory.


An incubator is a device that maintains a controlled environment for culturing animal cells. An incubator provides a constant temperature, humidity, carbon dioxide, and oxygen levels that are optimal for the growth and function of animal cells. An incubator also protects the cells from contamination by filtering the air and preventing temperature fluctuations. An incubator should be located in a stable and vibration-free area of the laboratory away from direct sunlight, heat sources, or air drafts. An incubator should be cleaned and disinfected regularly with 70% ethanol or another suitable disinfectant. An incubator should be operated according to the manufacturer's instructions and following the SOPs of the laboratory.


A microscope is an instrument that magnifies the image of small objects, such as animal cells, for observation and analysis. A microscope can be used for various purposes in animal cell culture, such as counting cells, assessing cell morphology, viability, and differentiation, detecting contamination, and performing live-cell imaging. The most common type of microscope used for animal cell culture is an inverted microscope, which has the light source and condenser above the specimen and the objective lens below it. This allows the observation of cells in culture vessels without disturbing them. An inverted microscope can be equipped with different features and accessories, such as phase contrast, fluorescence, digital camera, computer software, and micromanipulators, to enhance its performance and functionality.


A centrifuge is a device that spins a sample at high speed to separate its components based on their density and size. A centrifuge can be used for various purposes in animal cell culture, such as harvesting cells, washing cells, concentrating cells, separating cells from debris or supernatant, and isolating subcellular fractions. A centrifuge should be balanced properly before each run to prevent damage or injury. A centrifuge should be cleaned and disinfected regularly with 70% ethanol or another suitable disinfectant. A centrifuge should be operated according to the manufacturer's instructions and following the SOPs of the laboratory.


How to monitor and control the physical and chemical parameters of animal cell culture?




The physical and chemical parameters of animal cell culture are critical factors that affect the growth, survival, differentiation, and function of animal cells. These parameters include temperature, pH, dissolved oxygen, carbon dioxide, osmolality, and glucose. These parameters should be monitored and controlled regularly to ensure optimal conditions for animal cell culture.


Temperature is one of the most important parameters of animal cell culture, as it influences the metabolic rate, enzyme activity, membrane fluidity, and gene expression of animal cells. The optimal temperature for most mammalian cell cultures is 37C, which is close to the body temperature of humans and other warm-blooded animals. However, some cell types may require different temperatures depending on their origin or differentiation state. For example, some insect cell cultures require lower temperatures around 25-28C, while some stem cell cultures require higher temperatures around 39-40C. Temperature should be maintained within a narrow range of 0.5C to prevent thermal stress or shock that may damage or kill the cells.


pH is another important parameter of animal cell culture, as it affects the ionization state, charge distribution, and solubility of biomolecules and nutrients in the medium. The optimal pH for most mammalian cell cultures is around 7.2-7.4, which is close to the physiological pH of blood plasma. However, some cell types may require different pH values depending on their origin or differentiation state. For example, some tumor cell cultures require lower pH values around 6.5-6.8, while some alkalophilic cell cultures require higher pH values around 8.0-8.5. pH should be maintained within a narrow range of 0.1 units to prevent acidosis or alkalosis that may impair cellular functions.


Dissolved oxygen (DO) is another important parameter of animal cell culture, as it is essential for aerobic respiration and energy production of animal cells. The optimal DO level for most mammalian cell cultures is around 20-40%, which is close to the oxygen saturation level of air at sea level. However, some cell types may require different DO levels depending on their origin or differentiation state. For example, some hypoxic cell cultures require lower DO levels around 1-5%, while some hyperoxic cell cultures require higher DO levels around 60-80%. DO level should be monitored and controlled regularly to prevent hypoxia or hyperoxia that may affect cellular metabolism and gene expression.


Carbon dioxide (CO2) is another important parameter of animal cell culture, as it influences the pH and bicarbonate buffering system of the medium. The optimal CO2 level for most mammalian cell cultures is around 5%, which is close to the physiological CO2 level of blood plasma. However, some cell types may require different CO2 levels depending on their origin or differentiation state. For example, some anaerobic cell cultures require lower CO2 levels around 1-3%, while some photosynthetic cell cultures require higher CO2 levels around 10-15%. CO2 level should be maintained within a narrow range of 0.5% to prevent acid-base imbalance that may affect cellular functions.


Osmolality is another important parameter of animal cell culture, as it measures the concentration of solutes in the medium. The optimal osmolality for most mammalian cell cultures is around 280-320 mOsm/kg, which is close to the physiological osmolality of blood plasma. However, some cell types may require different osmolalities depending on their origin or differentiation state. For example, some marine cell cultures require higher osmolalities around 400-500 mOsm/kg, while some freshwater cell cultures require lower osmolalities around 200-250 mOsm/kg. Osmolality should be monitored and controlled regularly to prevent osmotic stress or shock that may cause cell swelling or shrinking.


Glucose is another important parameter of animal cell culture, as it is the main source of energy and carbon for animal cells. The optimal glucose level for most mammalian cell cultures is around 1-5 g/L, which is close to the physiological glucose level of blood plasma. However, some cell types may require different glucose levels depending on their origin or differentiation state. For example, some diabetic cell cultures require higher glucose levels around 10-15 g/L, while some ketogenic cell cultures require lower glucose levels around 0.5-1 g/L. Glucose level should be monitored and controlled regularly to prevent hyperglycemia or hypoglycemia that may affect cellular metabolism and function.


Animal Cell Culture Applications




Animal cell culture has a wide range of applications in biotechnology and medicine for various purposes, such as producing recombinant proteins, antibodies, viral vectors, tissues, organs, and drugs. Animal cell culture also provides a valuable tool for studying the structure, function, and interaction of animal cells, as well as their response to different stimuli and stressors. Some of the areas where animal cell culture has found most applications include cancer research, vaccine production, gene therapy, tissue engineering, and drug screening.


How to use animal cell culture for recombinant protein production and antibody engineering?




Recombinant protein production is the process of using genetically modified cells to produce proteins that are not normally expressed by them or are expressed at low levels. Antibody engineering is the process of using genetic engineering or other techniques to modify the structure and function of antibodies for specific purposes. Animal cell culture is widely used for both recombinant protein production and antibody engineering because it offers several advantages over other systems, such as bacteria, yeast, or plants. These advantages include:


  • The ability to perform post-translational modifications, such as glycosylation, phosphorylation, sulfation, and acylation, that are essential for the biological activity and stability of many proteins.



  • The ability to secrete proteins into the culture medium, which facilitates purification and recovery.



  • The ability to fold and assemble proteins correctly, which ensures proper function and reduces aggregation and degradation.



  • The ability to express complex proteins, such as antibodies, that require multiple subunits or domains.



Some examples of recombinant proteins produced by animal cell culture are insulin, erythropoietin, interferons, growth hormones, blood clotting factors, and vaccines. Some examples of antibodies engineered by animal cell culture are monoclonal antibodies, bispecific antibodies, antibody-drug conjugates, and chimeric antigen receptor (CAR) T cells.


How to use animal cell culture for viral vector production and gene therapy?




Viral vector production is the process of using genetically modified viruses to deliver genes or other molecules of interest into target cells or tissues. Gene therapy is the application of viral vector production to treat or prevent diseases by modifying the genes or gene expression of target cells or tissues. Animal cell culture is widely used for both viral vector production and gene therapy because it offers several advantages over other systems, such as bacteria, yeast, or plants. These advantages include:


  • The ability to produce high titers and quality of viral vectors that are compatible with human cells and tissues.



  • The ability to manipulate the genome and tropism of viral vectors to achieve specific and efficient gene delivery and expression.



  • The ability to test the safety and efficacy of viral vectors and gene therapy products in vitro and in vivo.



Some examples of viral vectors produced by animal cell culture are adenoviruses, adeno-associated viruses, retr


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