Experiment
JoVE Encyclopedia of Experiments
Encyclopedia of Experiments: Immunology
Encyclopedia of Experiments: Immunology
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Generating Recombinant Monoclonal Antibodies From Mammalian Cell Cultures
Transcript
Prepare the HEK cells in supplemented DMEM and grow them to 75% confluency in 75-square-centimeter culture flasks. For each heavy/light chain transfection, prepare a mix with 1.75 milliliters of DMEM without serum. Include 4.5 micrograms of heavy chain vector, 3 micrograms of light chain vector, and 50 microliters of polyethylenimine solution. Gently add this mixture to a plate of HEK cells and swirl the solution over the plate.
Incubate the plate overnight, and then begin collecting the culture media every day, replacing it with serum-less medium. The media contains the antibodies, and it can be collected daily for four days.
