HEK 293 cell suspension culture using fibronectin-adsorbed polymer nanospheres in serum-free medium.

Previously, we reported on suspension culture of anchorage-dependent animal cells using plain polymer nanospheres in serum-containing medium. For commercial cell culture, it is more advantageous to use serum-free medium than serum-containing medium. To culture anchorage-dependent animal cells using polymer nanospheres in serum-free medium, the nanospheres need to be coated with cell adhesion proteins. In this study, we utilized fibronectin-adsorbed polymer nanospheres for suspension culture of anchorage-dependent animal cells in serum-free medium. Fibronectin was adsorbed onto poly(lactic-co-glycolic acid) nanospheres (433 nm in average diameter) by immersing the nanospheres in fetal bovine serum. The nanospheres were used to culture human embryonic kidney (HEK) 293 cells in serum-free medium in stirred suspension bioreactors. Nanospheres attached between HEK 293 cells and promoted cell aggregate formation compared with culture without nanospheres. Most cells in the aggregates were viable over a 10-day culture period. Importantly, the use of poly(lactic-co-glycolic acid) nanospheres promoted the cell growth significantly, compared with culture without nanospheres (3.8- vs 1.8-fold growth). The nanosphere culture method developed in this study removes the time-consuming and costly process of adaptation of anchorage-dependent animal cells to suspension culture in serum-free medium. This culture method may be useful for the large-scale suspension culture of various types of anchorage-dependent animal cells in serum-free medium.