Lymphokine-activated Killer Cells: Culture Conditions for the Generation of Maximal in Vitro Cytotoxicity in Cells from Normal Donors1

The current method for generating lymphokine-activated killer ( I VK) cells for use in human clinical trials is both labor intensive and expensive. Therefore, we altered cell culture conditions to determine whether LAK cells with enhanced lytic activity could be generated. Culture of normal human peripheral blood leukocytes for 7 days generated LAK cells with 4-fold more lytic activity than culture for 3 days. Although cell viability over this 7-day period dropped from 94% on Day 3 to 73% by Day 7, the recovery of cells from culture increased from 61 to 106%. If cells were exposed to CO., lytic activity was further enhanced by up to 30-fold. Culture at a density of 1 or 2.5 x III"cells/ml caused no difference in cell viability, recovery, or LAK activity when cells were cultured for up to 4 days; however, when cells were cultured for longer times, an initial density of 1 x IO'1cells/ml yielded maximal LAK activity. Several commercially available serum-free defined media as well as human serum albumin supported LAK cell activation comparable to serum-containing media over a 4-day culture period. One defined medium, AIM V, supported LAK cell activation over a 7-day period even when cells were cultured at a density twice as high (2 x III'' cells/ml) as cells cultured in serumcontaining medium. The results demonstrate that simple manipulation of human LAK cell culture conditions generates cells with greatly enhanced lytic activity and that serum-containing medium may not be necessary for generating LAK cells under the current clinical protocols.