Membrane potential-sensitive fluorescent probes

Wolff, C. Fuks, B. Chatelain, P. Comparative study of membrane potential-sensitive fluorescent probes and their use in ion channel screening assays. J. Biomol. Screening 2003, 8, 533-543. 

Baxter DF, Kirk M, Garcia AF et al. (2002) A novel membrane potential-sensitive fluorescent dye improves cell-based assays for ion channels. J Biomol Screen 7(1) 79—85 Epps DE, Wolfe ML, Groppi V (1994) Characterization of the steady-state and dynamic fluorescence properties of the potential-sensitive dye bis-(l,3-dibutylbarbituric acid)trimethine oxonol (BiBAC4(3)) in model systems and cells. Chem Phys Lipids 69(2) 137—150 Gonzalez JE, Maher MP (2002) Cellular fluorescent indicators and voltage/ion probe reader (VIPR(TM)) tools for ion channel and receptor drug discovery. Recept Channels 8(5—6) 283—295... 

The use of potential-sensitive fluorescent probes to monitor the electrical potential across a cell membrane permits an accurate, noninvasive measurement of membrane potential changes in a wide variety of cells, vesicles, and organelles without the external electrical or mechanical manipulation required by micro-... 

Dragsten, P. R., and Webb, W. W., 1978, Mechanism of the membrane potential sensitivity of the fluorescent membrane probe mero-cyanine 540, Biochemistry 17 5228-5240. 

The membrane dipole potential ( d) can be measured using voltage-sensitive fluorescent probes. The use of di-8-ANEPPS for this purpose was first suggested by Loew and co-workers, who noted that the addition of 6-ketocholestanol, known to increase Wd, and phloretin, known to decrease fl d, to di-8-ANEPPS-bound man-branes caused a blue and a red shift, respectively, in the fluorescence excitation spectrum of the dye [13]. 

Pope, A. J. Leigh, R. A. Characterization of chloride transport at the tonoplast of higher plants using a chloride-sensitive fluorescent probe effects of other anions, membrane potential, and transport inhibitors. Planta 1990, 181, 406-413. 

Fluorescent, voltage-sensitive dyes can be employed to investigate the activity of ion channels by reflecting the cellular membrane potential (Epps et al. 1994 Plasek and Sigler 1996). This approach is suitable for use in conjunction with different measurement systems including Fluorometric imaging plate readers (FLIPR), Voltage/Ion probe readers (VIPR) or conventional fluorescence readers. 

Fluorescence is very sensitive to the chemical environment and may be utilized to provide information about the microenvironment surrounding the probe (5, 6). The fluorescence intensity (quantum yield) (1-3), the maximum emission wavelength (1-3), the fluorescence lifetime (6-9), or the polarization (10-12) may all be monitored for specific changes that are induced as a result of changes in polarity, pH, ion concentration, membrane potential, or ligand binding. 

The membrane or interfacial potential, particularly in biological applications, is often determined from the change in fluorescence of added carbocyanine dyes (12, 13). The fluorescence intensity of the dyes depends on the solvent in which the dyes are present. When the dyes are used as potential-sensitive probes, their fluorescent intensity is a function of the interfacial potential across the membrane. We studied the behavior of dye transport on a phase boundary between water and nitrobenzene to better understand the principles of the potential dye partitioning as a function of interfacial potential (141... 

Assays with externally added probes include the potential-sensitive dye safranin O (Figure 8). This cationic dye only loosely associates with an unpolarized vesicle membrane but binds more efficiently when an inside negative membrane potential is applied. This translocation into a more hydrophobic environment is accompanied with an increase in fluorescence and thereby reports on the extent of the applied membrane potential (Section A.A) ... 

Fluorescence is also a powerful tool for investigating the structure and dynamics of matter or living systems at a molecular or supramolecular level. Polymers, solutions of surfactants, solid surfaces, biological membranes, proteins, nucleic acids and living cells are well-known examples of systems in which estimates of local parameters such as polarity, fluidity, order, molecular mobility and electrical potential is possible by means of fluorescent molecules playing the role of probes. The latter can be intrinsic or introduced on purpose. The high sensitivity of fluo-rimetric methods in conjunction with the specificity of the response of probes to their microenvironment contribute towards the success of this approach. Another factor is the ability of probes to provide information on dynamics of fast phenomena and/or the structural parameters of the system under study. 

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