A comparison of techniques for introducing macromolecules into living cells.
Flow cytometry has been used to evaluate several techniques for introducing macromolecules into large numbers of living cells. One technique is cell fusion with red blood cell ghosts loaded with a fluorescent reporter molecule (RBCF). The second technique, termed osmotic lysis of pinosomes (OLP), involves a brief exposure of cells to a hypertonic solution containing the reporter molecule; subsequently, a hypotonic media is added which lyses the pinosomes formed during the hypertonic treatment. A third technique, scrape loading (SL), involves the creation of transient holes in the cell membrane through the application of mechanical forces, which allows for the passage of reporter molecules into cells. A comparison of these techniques is presented here. OLP appears to offer several advantages: It is a simple procedure, virtually all cells are fluorescently labelled, and it is capable of loading larger amounts of material more uniformly into cells while maintaining excellent viability.
Lee
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G
Department of Medicine, Columbia University, New York, New York 10032.
Delohery
T M
TM
Ronai
Z
Z
Brandt-Rauf
P W
PW
Pincus
M R
MR
Murphy
R B
RB
Weinstein
I B
IB
D002459:Cell Fusion; D003911:Dextrans; D004910:Erythrocyte Membrane; D004912:Erythrocytes; D005069:Evaluation Studies as Topic; D005434:Flow Cytometry / Q000379:methods; D005456:Fluorescent Dyes; D006982:Hypertonic Solutions; D008561:Membrane Fusion; D012235:Rhodamines
D003911:Dextrans; D005456:Fluorescent Dyes; D006982:Hypertonic Solutions; D012235:Rhodamines; C051311:rhodamine dextran