Filtroporation: A simple, reliable technique for transfection and macromolecular loading of cells in suspension

Cultured Chinese hamster ovary (CHO) cells suspended in their growth medium were forced by gas pressure through the uniformly sized micropores of filt er membranes. This procedure caused transient damage to the plasma membrane , which increased the permeability of the cells to exogenous molecules. Thi s "filtroporation" was indicated by uptake of fluorescent dextran molecules up to 500,000 MW in cells deemed viable by trypan blue dye exclusion. The macromolecular uptake was increased if the driving pressure was increased a t constant micropore size, or if the micropore size was decreased at consta nt driving pressure. Larger membrane perturbations permitted uptake of a lu ciferase reporter plasmid, which resulted in transfection of the CHO cells with the surviving cells expressing luciferase activity after 2 days in cul ture. This simple and general new method of porating cells in suspension ma y be optimized to incorporate the desired macromolecules white retaining th e maximum viability. (C) 1999 John Wiley & Sons, Inc.