Culture of animal cells pptx

[...]... the recovery of total cells per embryo, cells/ g, viable cells/ embryo, and viable cells/ g 4) From the number of cells recovered at the first subculture (3 days in this case), calculate the yield of cells per embryo, and as a ratio of the total cells seeded, and viable cells seeded 5) Was the dye exclusion staining a good predictor of recovery? Exercise 21 Cloning of Monolayer Cells Purpose of Procedure... seeded n Sample Cells/ flask at seeding c×d×v×r×f c×d×v×r×f n 200,000 30642000 153.21 Ancillary Protocol: Scale-up in Suspension (see Protocol 26.1) Example Experimental Variations Comparison of subculture from log-phase and plateauphase cells Background: Cell Section 13.7.3) concentration at subculture (see Log-phase cells Plateau-phase cells 22 CULTURE OF ANIMAL CELLS Exercise 14 Subculture of Continuous... TABLE 1.5 Subculture Advantages Disadvantages Propagation Trauma of enzymatic or mechanical disaggregation Selection of cells adapted to culture Overgrowth of unspecialized or stromal cells Genetic instability Loss of differentiated properties (may be inducible) Increased risk of misidentification or cross-contamination More cells Possibility of cloning Increased homogeneity Characterization of replicate... cells by pipetting vigorously to give a single cell suspension vi) Count cells by hemocytometer or electronic cell counter vii) Calculate number of cells per flask, cells/ mL culture medium, and cells/ cm2 at time of trypsinization d) Fix and stain cells in other flasks (see Protocol 16.2) Note: Pipettors should only be used for counting cells from isolated samples and not for dispensing cells for subculture,... cell morphology of fed and unfed cultures 2) Photograph before (phase-contrast illumination) and after (normal bright-field illumination) staining Exercise 16 Curve Construction and Analysis of Growth Purpose of Procedure Familiarization with the pattern of regrowth following subculture; demonstration of the growth cycle in routine subculture and as an analytical tool 24 CULTURE OF ANIMAL CELLS Applications... 2.1 are regarded as essential 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 Ian Freshney Copyright  2005 John Wiley & Sons, Inc 11 12 CULTURE OF ANIMAL CELLS TABLE 2.1 Training Programs Exercise... Pipetting and transfer of fluids Observation of cultured cells 3 Aseptic technique: preparing medium for use 4 5 Feeding a culture Washing and sterilizing glassware 6 Preparation and sterilization of water 7 Preparation of PBS 8 Preparation of pH standards 9 Preparation of stock medium and sterilization by filtration Preparation of complete medium from powder or 10× stock Counting cells by hemocytometer... Monolayer phase-contrast microscopy A demonstration of trypsinization (see Protocol 13.2) will be required Exercise Purpose of Procedure Propagating a culture by transferring the cells of a culture to a new culture vessel This may involve dilution to reseed the same size of culture vessel, or increasing the size of vessel if expansion is required Summary of Procedure A cell monolayer is disaggregated in... Quantitative skill Counting cells and assessment of viability Evaluation of relative merits of two methods Assessing a culture Aseptic handling Cell counting and viability Selecting reseeding concentration Assessing a culture Aseptic handling How to disaggregate cells Technique of trypsinization Cytology of cells Phase-contrast microscopy Fixation and staining Photography Replicate subcultures in multiwell... is an acceptable level of error in the precision of pipetting? 4) Which is more important: absolute accuracy or consistency? Exercise 2 Introduction to Cell Cultures Purpose of Procedure Critical examination of cell cultures Applications Checking consistency during routine maintenance; evaluation of status of cultures before feeding, subculture, or cryopreservation; assessment of response to new or . Growth of Tissue Culture. Number of hits in PubMed for ‘‘cell culture ’ from 1965. The pre-1960 figure is derived from the bibliography of Fischer [1925]. Culture of Animal Cells: A Manual of Basic. them attractive systems for the study of the molecular basis of development. More recently, the needs of agriculture and pest control have 4 CULTURE OF ANIMAL CELLS encouraged toxicity and virological. 1.5. Subculture Advantages Disadvantages Propagation Trauma of enzymatic or mechanical disaggregation More cells Selection of cells adapted to culture Possibility of cloning Overgrowth of unspecialized or