Quantitative introduction of a given macromolecule into cells by fusion with erythrocyte ghosts using a fluorescence activated cell sorter.

Abstract

FITC-conjugated bovine serum albumin (FITC-BSA) molecules were quantitatively introduced into human erythrocyte ghosts by gradual hemolysis. When the ghosts and L cells were fused with UV-inactivated HVJ (Sendai virus), FITC-BSA was introduced into the cytoplasm of the L cells and fluorescence could be observed inthe cells with a fluorescence microscope. A mixture of L cells and ghosts was introduced into a fluorescence activated cell sorter (FACS), which could separate the mononuclear cells on the basis of their light-scattering profile. Four distinct populations of mononuclear cells were found by fluorescence analysis. These populations were separated from the cell mixture and found to correspond to cells fused with one, two and three ghosts and unfused cells. After separation, the cells from each population could form colonies in culture. As a given macromolecule can be quantitatively introduced into erythrocyte ghosts with the FITC-BSA, after fusion of these ghosts with cells, this sorting method is useful for separating cells containing a definite number of macromolecules.