Preparation of Mammary Epithelial Cells from Reduction Mammoplasty Specimens 22. Primary Culture of Human Astrocytes, 424 Protocol 22. This recognition may mark the transition from an era of fundamental molecular biology, in which many of the regulatory processes have been worked out at the cellular level, to an era of cell or tissue biology, in which that understanding is applied to integrated populations of cells and to a more precise elaboration of the signals transmitted among cells. The work area, in its simplest form, should be a plastic laminate-topped bench, preferably plain white or neutral gray, to facilitate the observation of cultures, dissection, etc. Too Many Colonies per Dish, 587 29. Experimental Variations Venting flasks 1 Prepare medium according to Protocol 11. In tropical regions, or where overheating is a frequent problem, it may be necessary to incorporate cooling coils in the duct work of the heaters.
Increased understanding of the constitution of medium and its supplementation. Cultures of Neuronal Aggregates, 492 Protocol 25. Growth on Confluent Feeder Layers 24. Exercise Summary of Procedure There are two options for this exercise: a simple growth curve of one cell line using flasks as for regular subculture, or using multiwell plates to analyze differences in growth at different densities, between two different cell lines, or under any other selected set of conditions. Chairs should be a suitable height, with adjustable seat height and back angle, and able to be drawn up close enough to the front edge of the hood to allow comfortable working well within it. Demonstration material or operations: Use of osmometer or conductivity meter.Next
Incubators also lose more heat when they are opened and are slower to recover than a hot room. For this reason, many workers have attempted to reconstitute three-dimensional cellular structures by using aggregates in cell suspension see Section 25. The book includes a comprehensive list of suppliers for all equipment used in the protocols presented, with websites available in an appendix. Safety: Care should be taken when handling human cells see Exercise 10 and Section 7. Cell Cycle Inhibition and Progression. When these are inactivated, usually by phosphorylation, cells proceed round the cycle b. Preparation and Sterilization, 50 4.Next
These Culture of Animal Cells: A Manual of Basic Technique, Fifth Edition, by R. The matrix adheres to the charged substrate, and the cells then bind to the matrix via specific receptors. Disadvantages of Serum-Free Media, 122 9. Both require a clear concise style, but the author has more freedom to express ideas and opinions when writing a book. Training objectives Isolation of cell type with desired phenotype by one of several separation methods.Next
Confluent Feeder Layers, 466 Protocol 24. For general information on our other products and services or for technical support, please contact our Customer Care Department within the United States at 800 762-2974, outside the United States at 317 572-3993 or fax 317 572-4002. The term transformation has been applied to the process of formation of a continuous cell line partly because the culture undergoes morphological and kinetic alterations, but also because the formation of a continuous cell line is often accompanied by an increase in tumorigenicity. As the maintenance and proliferation of specialized cells, and the induction of their differentiation, may depend on paracrine and endocrine factors, these must be identified and added to differentiation medium see Sections 17. The installation will pay for itself eventually in the cost of cylinders of mixed gases for gassing cultures, and it provides a 47 better supply, which can be protected see Section 5. A tour of the tissue culture facilities is an essential introduction; this lets the trainee meet other staff, determine their roles and responsibilities, and see the level of preparation Culture of Animal Cells: A Manual of Basic Technique, Fifth Edition, by R. Neither the publisher nor author shall be liable for any loss of profit or any other commercial damages, including but not limited to special, incidental, consequential, or other damages.
Autoclaves, ovens, and distillation apparatus should be located in a separate room if possible see Figs. The trainee is then required to determine the cell concentration and select the correct concentration for reseeding, instilling a concept of quantitation in cell culture and enhancing numeracy skills. It is during G1 that the cell is particularly susceptible to control of cell cycle progression at a number of restriction points, which determine whether the cell will re-enter the cycle, withdraw from it, or withdraw and differentiate. Authentication and Validation, 259 14. The cell cycle is arrested at restriction points or checkpoints by the action of Rb, p53, and other cell cycle inhibitors a.Next
It is assumed that the inside of a pipettor is sterile or does not displace enough air for this to matter. Estimation of Viability by Dye Uptake, 367 21. Subculture of Monolayer Cells 12. When a new building is contemplated, there is more scope for integrated and innovative design, and facilities may be positioned for ergonomic and energy-saving reasons, rather than structural ones. Reusable Sterilizing Filters, 139 Protocol 10. Liquid-Nitrogen Store and Cryostore 4. The combined use of both counting methods will be incorporated in the following description.Next
Demonstration material or operations: Trainee should observe all steps in preparation and participate where possible; this may require repeated short visits to see all procedures. In many respects, it is better to provide individual peristaltic pumps at each workstation see Figs. Standard Protocol Subculture in Suspension see Protocol 13. This text is an indispensable resource for those in or entering the field, including academic research scientists, clinical and biopharmaceutical researchers, undergraduate and graduate students, cell and molecular biology and genetics lab managers, trainees and technicians. Exercise Summary of Procedure Examine and photograph a range of cell lines at different cell densities. Ancillary Protocols: Preparation of Conditioned Medium see Protocol 14. Tissue Disaggregation in Warm Trypsin, 173 11.Next