Introduction of macromolecules into cultured mammalian cells by osmotic lysis of pinocytic vesicles

Abstract

We have developed a new procedure for introducing macromolecules into cultured mammalian cells based on osmotic lysis of pinocytic vesicles. Cells are first incubated in culture medium containing 0.5 M sucrose, 10% polyethylene glycol 1000 and the macromolecule to be transferred. Cells are then placed in medium diluted with 0.66 parts water. Most pinocytic vesicles formed in the presence of sucrose burst in hypotonic medium, thereby releasing the enclosed macromolecule. L929 cells remain fully viable after a single hypertonic sucrose treatment, and a majority survives four successive rounds of osmotic lysis. This procedure, termed osmotic lysis of pinosomes, has been used to transfer substantial amounts of horseradish peroxidase, antiricin antibodies and dextran 70,000 into the cytosol of L929 cells. Direct comparison of the degree of ricin resistance conferred by transfer of antiricin antibodies revealed pinosome lysis to be equal, if not superior, to injection mediated by red blood cells.