Fluorometric assay using dimeric dyes for double- and single-stranded DNA and RNA with picogram sensitivity.

摘要: Thiazole orange homodimer (TOTO; 1,1′-(4,4,7,7-tetramethyl-4,7-diazaundecamethylene)-bis-4-[3-methyl-2,3-dihydro-(benzo-1,3-thiazole)-2-methylidene]-quinolinium tetraiodide) and oxazole yellow (YOYO; an analogue of TOTO with a benzo-1,3-oxazole in place the benzo-1,3-thiazole) bind very high affinity to nucleic acids more than 1000-fold fluorescence enhancement upon binding. A linear dependence intensity on DNA concentration over range from 0.5 100 ng/ml presence 2 × 10−7 M or YOYO 4 mM Tris-acetate/0.1 EDTA/50 NaCl, pH 8.2 allows sensitive quantitation double-stranded conventional fluorometer. With acid-dye mixtures array 25-μl wells block low autofluorescence plastic detection laser-excited confocal scanner, as little 20 pg can be detected per well. The scanning method is rapid, has throughput, requires small amounts sample. It also single-stranded RNA.