Selective staining of biomembranes using voltage-sensitive dyes

A method for the enzymatic activation of an amphiphilic voltage-sensitive dye comprising

• (a) providing a precursor to the amphiphilic voltage-sensitive dye with said precursor having an enzymatically cleavable hydrophilic group at a lipophilic tail of the amphiphilic voltage-sensitive dye,
• (b) cleaving the precursor to the amphiphilic voltage-sensitive dye using an enzyme expressed by a target cell, thereby forming the amphiphilic voltage-sensitive dye, and
• (c) binding the amphiphilic voltage-sensitive dye to a lipophilic interior of a lipid membrane of the target cell, wherein the enzyme is overexpressed by a transfected cell and provided with a targeting signal which directs the enzyme to the cell wall.

A method for the enzymatic activation of an amphiphilic voltage-sensitive dye comprising

• (a) providing a precursor to the amphiphilic voltage-sensitive dye with said precursor having an enzymatically cleavable hydrophilic group at a lipophilic tail of the amphiphilic voltage-sensitive dye,
• (b) cleaving the precursor to the amphiphilic voltage-sensitive dye using an enzyme expressed by a target cell, thereby forming the amphiphilic voltage-sensitive dye, and
• (c) binding the amphiphilic voltage-sensitive dye to a lipophilic interior of a lipid membrane of the target cell, wherein dye not bound to the membrane is reconverted into the dye precursor by a second enzyme.

A voltage sensitive dye precursor having the formula (I)

• X-A-Z-R1R2 (I)
• wherein A denotes a voltage sensitive chromophore selected from
• X denotes a polar head group selected from —(CH2)m—SO2H, —(CH2)m—N+(CH3)3 or —CH2—CHOH—CH2—N+(CH3)2—CH2—CH2—OH, m is an integer from 0 to 6, Z is selected from N, P, As, Sb or Bi and R1 and R2 independently at each occurrence represent a C10-C20 hydrocarbon residue which is substituted with at least one group selected from a (poly-)phosphate group or a (poly-)carbohydrate group.