Lab 7&8 - Cell culture and quantification

Lab 7&8 - Cell culture and quantification -...

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Bioengineering 167L Labs 7 & 8 Mammalian Cell Culture Mammalian cell culture is one of the most essential tools in the areas of biotechnology and cell biology. In this lab, you will learn how to culture a widely used cell line, the HeLa cells, and learn ways of quantifying the cells. Required Equipment: Biosafety cabinet, CO 2 incubator, Pipet-aid Chemicals: Complete MEM media (from lab 6) PBS (from lab 6) Trypsin (In this lab, students will use a synethic Trypsin, called TrypLE™ Express, instead) Supplies: T75 culture flasks, 15 ml tubes, hemocytometer SAFETY: Wear gloves, safety glasses, and lab coats. Basic outline for lab 7 and 8 Laboratory 7 o TA’s demo Cell passaging demo o Students will passage cells at different split ratios - see Details of Laboratory 7: Passaging Cells There will be a flask ready for each student . Use the complete media and phosphate-buffered saline (PBS) made from previous lab. Students will label and use the same media and PBS for the next laboratory meeting. Students will passage cells with different split ratios. Per group: o One student will split his/her flask in ratios of 1:10 and 2:10. o The other student will use passage a new flask of cells into another flask using a split ratio of 3:10. Use a hemocytometer to determine the cell concentration in the original cell suspension. o Cleaning up and dealing with the biohazard waste. For the solutions, bleach until each solution reaches approximately 10 volume% bleach, wait for 10 minutes, and dump the solution down the drain with running water. Solid wastes will be taken by the TA to the autoclave site.
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Laboratory 8 o The details of this laboratory can be found in – see Details of Laboratory 8. o Per group: With the cells in the three flasks (i.e. those resulting from the 1:10, 2:10, and 3:10 split ratios), conduct the following: Count the cells using the hemocytometer. o
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Lab 7&8 - Cell culture and quantification -...

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