Introduction to Cell Culture Biotechnology I

Cell Culture Definition: the in vitro growth of cells isolated from multi-cellular organisms Process: Cells will continue dividing until they fill up the container; cell to cell contact stops cell division Uses: vaccines, research of all kinds including stem cell, recombinant DNA, production of antibodies

Types of Cell used Mammalian cells are commonly used but they do grow slower and are more fragile than bacterial cells. Also have more complex nutrient requirements Bacterial cells were first used to make genetically engineered products and they reproduce very quickly

Growing Bacterial Cells Choose bacteria – E. coli most common Make media Petri plates use agar in media (Luria Broth, nutrient agar) Liquid cultures use broth (LB, nutrient broth) Sterilize media in autoclave Pour media plates Innoculate media Grow cells in incubator (37oC)

Equipment Required for Mammalian Cells CO2 incubator: for proper atmospheric conditions Laminar flow hood: for sterility Hemocytometer: for cell counting Inverted Microscope: for viewing cells on hemocytometer

Cell Culture Process Contaminating the cell culture is area of concern To avoid contamination mfg. techs do the following: Sterilize all containers used Change the media frequently Media is the source of nutrition, pH indicator often times included in media to help monitor pH which is critical to cell survival

Cell Culture Process Count cells Using a hemocytometer the cells are counted to determine if they are ready to harvest Can stain cells with trypan blue to determine if they are still alive. All cells which exclude the dye are viable. All stained cells are dead. Live cell Dead cell

Cell Culture Process View cells Inverted microscope is used to determine cell growth and activity Adherent cells are ones which are stuck to the flask or vessel Cell counting video for cell culture