Nuclear organization studied with the help of a hypotonic shift: its use permits hydrophilic molecules to enter into living cells

A new procedure for introduction of hydrophilic molecules into living cells based on efficient uptake of these molecules into the cells during hypoton ic treatment is presented and its use is demonstrated by a variety of appli cations. Experiments with cultured vertebrate and Drosophila cells and vari ous animal tissues demonstrated that the increase in cell membrane permeabi lity under hypotonic conditions is a general phenomenon in all animal cells tested. The efficiency of the method depends on the composition and temper ature of the hypotonic buffer, the duration of the hypotonic treatment and the molecular weight of the molecules introduced into living cells. The ver satility of this approach is demonstrated with various types of molecules s uch as modified nucleotides, nucleotides with conjugated fluorochrome, pept ides, phosphatase substrates and fluorescent dyes. The method opens new pos sibilities for the direct investigation of a variety of biological problems as documented here with data on the functional organization of the cell nu cleus.