By Shirley McBride, Mary Heenan, Martin Clynes (auth.), Dr. Martin Clynes (eds.)
Cell tradition options permit various molecular and mobilephone organic inquiries to be addressed, providing physiological stipulations when keeping off using laboratory animals. as well as easy options, quite a lot of specialized useful protocols masking the subsequent components are integrated: mobilephone proliferation and loss of life, in-vitro types for mobile differentiation, in-vitro types for toxicology and pharmacology, commercial program of animal phone tradition, genetic manipulation and research of human and animal cells in culture.
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Extra resources for Animal Cell Culture Techniques
Slow growing cells will have a low split ratio. 2. 1. 2. Split ratio Cell type Split ratio African green monkey kidney (Vero) Fetal rhesus kidney (FRK) Human epithelium (Hep-2) Human embryonic lung (MRC-5) Human rhabdomyosarcoma (RD) Madin Darby canine kidney (MDCK) 1:3 1:2 1:5 1:2 1:2 1:4 2 Cell Culture and Diagnostic Virology 21 Anchorage-dependent cells are dispersed from the monolayers using equal volumes (10 ml) of a chelating agent, versene (EDTA), and a proteolytic enzyme, trypsin. Cells may also be removed by scraping with a disposable cell scraper or by using sterile glass beads.
Note: Check pH perodically, as it is critical for optimal fluorescence of the DNA stain. 3 Screening for Mycoplasma Contamination in Animal Cell Cultures 43 Preparation of (overslips and Slides 1. Soak the required number of coverslips and slides in detergent for 1 h. 2. Rinse thoroughly in tap water for 5 min and in deionized water for a further 5 min. 3. Rinse the coverslips in methanol and wipe dry with lint-free tissue. The slides can be refrigerated in methanol until required (use a sparkproof refrigerator).
Examine the condition of the cell monolayer using an inverted micro- scope and ensure that the cells are healthy and confluent. 2. Discard the spent growth medium (GM) from the vessel and wash the monolayer twice with 10 ml of prewarmed versene/trypsin wash solution leaving the solution on the monolayer for 20 s with each wash. 3. Discard the wash solution and incubate the flask at 37°C for 2-5 min. Some cells are difficult to detach from the monolayer and require a Fig. 2. Selection of microtiterplates used in cell culture subculturing procedure 22 EDWIN O'KELLY more vigorous removal routine.
Animal Cell Culture Techniques by Shirley McBride, Mary Heenan, Martin Clynes (auth.), Dr. Martin Clynes (eds.)