Long-wavelength iodide-sensitive fluorescent indicators for measurement of functional CFTR expression in cells

1 Departments of Medicine and Physiology, Cardiovascular Research Institute, University of California, San Francisco, California, 94143-0521; and 2 Faculty of Chemistry, A. Mickiewicz University, 60-780 Poznan, Poland Limitations of available indicators [such as 6-methoxy- N -(3-sulfopropyl)quinolinium (SPQ)] for measurement of intracellular Cl are their relatively dim fluorescence and need for ultraviolet excitation. A series of long-wavelength polar fluorophores was screened to identify compounds with Cl and/or I sensitivity, bright fluorescence, low toxicity, uniform loading of cytoplasm with minimal leakage, and chemical stability in cells. The best compound found was 7-( - D -ribofuranosylamino)-pyrido[2,1-h]-pteridin-11-ium-5-olate (LZQ). LZQ is brightly fluorescent with excitation and emission maxima at 400-470 and 490-560 nm, molar extinction 11,100 M 1 · cm 1 (424 nm), and quantum yield 0.53. LZQ fluorescence is quenched by I by a collisional mechanism (Stern-Volmer constant 60 M 1 ) and is not affected by other halides, nitrate, cations, or pH changes (pH 5-8). After LZQ loading into cytoplasm by hypotonic shock or overnight incubation, LZQ remained trapped in cells (leakage