This Article Full Text (PDF) References Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of the journal Add to Saved Citations Download to citation manager Request Permissions Request Reprints Add to My Marked Citations Citing Articles Citing Articles via HighWire Citing Articles via Web of Science (56) Citing Articles via Google Scholar Citing Articles via Scopus Google Scholar Articles by Baxter, D. F. Articles by Xie, Y. Search for Related Content PubMed PubMed Citation Articles by Baxter, D. F. Articles by Xie, Y. Pubmed/NCBI databases Gene GEO Profiles HomoloGene Protein UniGene Compound via MeSH Substance via MeSH Hazardous Substances DB POTASSIUM POTASSIUM CHLORIDE Social Bookmarking                         What's this?

A Novel Membrane Potential-Sensitive Fluorescent Dye Improves Cell-Based Assays for Ion Channels

Millennium Pharmaceuticals, Inc., Cambridge, MA

Martin Kirk

Molecular Devices Corporation, Sunnyvale, CA, Celtor Biosystems, Santa Clara, CA

Amy F. Garcia

Millennium Pharmaceuticals, Inc., Cambridge, MA

Alejandra Raimondi

Millennium Pharmaceuticals, Inc., Cambridge, MA

Mats H. Holmqvist

Millennium Pharmaceuticals, Inc., Cambridge, MA

Kimberly K. Flint

Millennium Pharmaceuticals, Inc., Cambridge, MA

Dejan Bojanic

Millennium Pharmaceuticals, Inc., Cambridge, MA

Peter S. Distefano

Millennium Pharmaceuticals, Inc., Cambridge, MA, Elixir Pharmaceuticals, Inc., Cambridge, MA

Rory Curtis

Millennium Pharmaceuticals, Inc., Cambridge, MA, Elixir Pharmaceuticals, Inc., Cambridge, MA

Yu Xie

Millennium Pharmaceuticals, Inc., Cambridge, MA

The study of ion channel-mediated changes in membrane potential using the conventional bisoxonol fluorescent dye DiBAC₄(3) has several limitations, including a slow onset of response and multistep preparation, that limit both the fidelity of the results and the throughput of membrane potential assays. Here, we report the characterization of the FLIPR Membrane Potential Assay Kit (FMP) in cells expressing voltage- and ligand-gated ion channels. The steady-state and kinetics fluorescence properties of FMP were compared with those of DiBAC₄(3), using both FLIPR and whole-cell patch-clamp recording. Our experiments with the voltage-gated K⁺ channel, hElk-1, revealed that FMP was 14-fold faster than DiBAC₄(3) in response to depolarization. On addition of 60 mM KCl, the kinetics of fluorescence changes of FMP using FLIPR were identical to those observed in the electrophysiological studies using whole-cell current clamp. In addition, KCl concentration-dependent increases in FMP fluorescence correlated with the changes of membrane potential recorded in whole-cell patch clamp. In studies examining vanilloid receptor-1, a ligand-gated nonselective cation channel, FMP was superior to DiBAC₄(3) with respect to both kinetics and amplitude of capsaicin-induced fluorescence changes. FMP has also been used to measure the activation of K_ATP¹ and hERG.² Thus this novel membrane potential dye represents a powerful tool for developing high-throughput screening assays for ion channels.

Journal of Biomolecular Screening, Vol. 7, No. 1, 79-85 (2002)
DOI: 10.1177/108705710200700110


CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    Twitter    What's this?

This article has been cited by other articles:

				N. Y. Yount, D. Kupferwasser, A. Spisni, S. M. Dutz, Z. H. Ramjan, S. Sharma, A. J. Waring, and M. R. Yeaman Selective reciprocity in antimicrobial activity versus cytotoxicity of hBD-2 and crotamine PNAS, September 1, 2009; 106(35): 14972 - 14977. [Abstract] [Full Text] [PDF]

				J. George, S. M. Dravid, A. Prakash, J. Xie, J. Peterson, S. V. Jabba, D. G. Baden, and T. F. Murray Sodium Channel Activation Augments NMDA Receptor Function and Promotes Neurite Outgrowth in Immature Cerebrocortical Neurons J. Neurosci., March 11, 2009; 29(10): 3288 - 3301. [Abstract] [Full Text] [PDF]

				C. Joesch, E. Guevarra, S. P. Parel, A. Bergner, P. Zbinden, D. Konrad, and H. Albrecht Use of FLIPR Membrane Potential Dyes for Validation of High-Throughput Screening with the FLIPR and {micro}ARCS Technologies: Identification of Ion Channel Modulators Acting on the GABAA Receptor J Biomol Screen, March 1, 2008; 13(3): 218 - 228. [Abstract] [PDF]

				E. F. Vernon-Wilson, F. Aurade, L. Tian, I. C. M. Rowe, M. J. Shipston, J. Savill, and S. B. Brown CD31 delays phagocyte membrane repolarization to promote efficient binding of apoptotic cells J. Leukoc. Biol., November 1, 2007; 82(5): 1278 - 1288. [Abstract] [Full Text] [PDF]

				B. D. Guth Preclinical Cardiovascular Risk Assessment in Modern Drug Development Toxicol. Sci., May 1, 2007; 97(1): 4 - 20. [Full Text] [PDF]

				K. J. Cassutt, M. J. Orsini, M. Abousleiman, D. Colone, and W. Tang Identifying Nonselective Hits from a Homogeneous Calcium Assay Screen J Biomol Screen, March 1, 2007; 12(2): 285 - 287. [Abstract] [PDF]

				M. Spitzner, J. Ousingsawat, K. Scheidt, K. Kunzelmann, and R. Schreiber Voltage-gated K+ channels support proliferation of colonic carcinoma cells FASEB J, January 1, 2007; 21(1): 35 - 44. [Abstract] [Full Text] [PDF]

				W. K. Kroeze and B. L. Roth Screening the receptorome J Psychopharmacol, July 1, 2006; 20(4_suppl): 41 - 46. [Abstract] [PDF]

				D. G. Nicholls Simultaneous Monitoring of Ionophore- and Inhibitor-mediated Plasma and Mitochondrial Membrane Potential Changes in Cultured Neurons J. Biol. Chem., May 26, 2006; 281(21): 14864 - 14874. [Abstract] [Full Text] [PDF]

				E. R. Benjamin, F. Pruthi, S. Olanrewaju, V. I. Ilyin, G. Crumley, E. Kutlina, K. J. Valenzano, and R. M. Woodward State-Dependent Compound Inhibition of Nav1.2 Sodium Channels Using the FLIPR Vm Dye: On-Target and Off-Target Effects of Diverse Pharmacological Agents J Biomol Screen, February 1, 2006; 11(1): 29 - 39. [Abstract] [PDF]

				A. Dorn, F. Hermann, A. Ebneth, H. Bothmann, G. Trube, K. Christensen, and C. Apfel Evaluation of a High-Throughput Fluorescence Assay Method for hERG Potassium Channel Inhibition J Biomol Screen, June 1, 2005; 10(4): 339 - 347. [Abstract] [PDF]

				M. Estacion, S. Li, W. G. Sinkins, M. Gosling, P. Bahra, C. Poll, J. Westwick, and W. P. Schilling Activation of Human TRPC6 Channels by Receptor Stimulation J. Biol. Chem., May 21, 2004; 279(21): 22047 - 22056. [Abstract] [Full Text] [PDF]

				C. Wolff, B. Fuks, and P. Chatelain Comparative Study of Membrane Potential-Sensitive Fluorescent Probes and their Use in Ion Channel Screening Assays J Biomol Screen, October 1, 2003; 8(5): 533 - 543. [Abstract] [PDF]

				R. W. Fitch, Y. Xiao, K. J. Kellar, and J. W. Daly Membrane potential fluorescence: A rapid and highly sensitive assay for nicotinic receptor channel function PNAS, April 15, 2003; 100(8): 4909 - 4914. [Abstract] [Full Text] [PDF]