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

Bacterial cells were first used to make genetically engineered products and they reproduce very quickly

Mammalian cells are commonly used but they do grow slower and are more fragile than bacterial cells. Also have more complex nutrient requirements

Culturing Bacteria

Use Sterile Technique Make media

E. coli grows best in Luria Broth Agar for plates and broth for liquid cultures

Sterilize media in autoclave Pour plates Innoculate plates and/or broth Incubate plates or broth in 37oC incubator

Equipment Required for Other 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

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

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.

Dead cellLive cell