Encyclopedia of Experiments: Immunology
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On the day of transfection, prepare liposome and DNA solutions per the manufacturer's recommendations. Transfect the DNA cells with a DNA composition of 1:1:2 of HA:NA:Gag and a total DNA quantity of 40 micrograms per T-150 flask. Repeat this step to create nine T-150 flasks for a total volume of 200 milliliters.
Next, dilute the DNA and liposome solutions in serum-free transfection media without antibiotics so that each flask contains a total volume of 24 milliliters. Return the flasks to the incubator and maintain the cells in transfection culture medium until the day of virus-like particle or VLP, harvest.
Transfer the supernatant into 15-milliliter conical tubes to harvest the culture from the cells after 72 to 96 hours post-transfection. Spin the cells down to pellet the cellular debris. Collect the supernatants and filter them through a 0.22-micron pore membrane.
