Introduction of definite amounts of nonpermeant molecules into living cells after electropermeabilization: direct access to the cytosol

Exp Cell Res. 1988 Mar;175(1):15-25. doi: 10.1016/0014-4827(88)90251-0.

Abstract

The possibility of introducing definite amounts of nonpermeant molecules into electropermeabilized living cells has been approached by quantifying the amounts of Lucifer Yellow (LY; a 457-Da highly fluorescent molecule) and Phytolacca americana (Pokeweed) antiviral protein (PAP; a 30,000-Da ribosome-inactivating protein) retained by the cells after closure of the electric-field-induced transient structures of permeation. Without the electropermeabilization, these two molecules enter the cell only in very small amounts by fluid-phase pinocytosis. Under our experimental conditions, using the NIH 3T3 cells, the intracellular LY concentration can reach a value equivalent to the extracellular concentration and can be regulated by controlling the external concentration. We describe the use of LY in a rapid and efficient test for the determination of the best electrical-shock conditions of other cell lines. After electropermeabilization, PAP is 2 X 10(5) times more cytotoxic. Its toxicity can be detected at external concentrations (10(-11) M) corresponding to less than 10 internalized molecules per electropermeabilized cell. Therefore, after electropermeabilization, the nonpermeant molecules have a direct access to the cytosol and the biological effect of nonpermeant substances can be revealed.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Line
  • Cell Membrane Permeability*
  • Cell Survival
  • Cytosol
  • Electricity*
  • Humans
  • In Vitro Techniques
  • Isoquinolines*
  • Membrane Potentials
  • Mice
  • Molecular Weight
  • N-Glycosyl Hydrolases*
  • Plant Proteins*
  • Ribosome Inactivating Proteins, Type 1

Substances

  • Isoquinolines
  • Plant Proteins
  • Ribosome Inactivating Proteins, Type 1
  • lucifer yellow
  • N-Glycosyl Hydrolases
  • pokeweed antiviral protein