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Free keywords:
PLACENTAL-ALKALINE-PHOSPHATASE; ANELLATED HEMICYANINE DYES;
MEMBRANE-POTENTIAL CHANGES; CHARGE-SHIFT PROBES; FLUORESCENT DYES;
MAMMALIAN-CELLS; NEURON MEMBRANE; PROTEIN; ABSORPTION; DYNAMICS
Abstract:
Optical recording of action potentials in individual neurons requires
cell-selective targeting with a fluorescent, voltage-sensitive probe. We
report on a new labeling system that takes advantage of recent
developments in prodrug-based chemistry and allows for the targeting of
a lipophilic dye into the plasma membrane of genetically specified
cells. With the introduction of two phosphonooxymethylammonium
zwitterions into the hydrocarbon chains of an amphiphilic,
voltage-sensitive hemicyanine dye, a precursor dye was formed that is
water-soluble to an extent that it can no longer bind into cell
membranes and hence prevents unspecific staining.
Glycosylphosphatidylinositol anchored placental alkaline phosphatase
expressed in HEK293 cells converted the precursor dye to a homologue of
the widely used dye di-4-ANEPPS and gave rise to excellent levels of
plasma membrane localized staining. The voltage sensitivity of the
enzymatically activated dye was tested and shown to be similar to
sensitivity reported for di-4-ANEPPS.
