Abstract: Methods of determining the presence or amount of a target polynucleotide in a sample are provided. A sample that contains a target polynucleotide, a polynucleotide that may include nucleic acid analogs and that is complementary to a target nucleic acid sequence of the target polynucleotide, and a dye for which the rate of change in an optical property is different in the presence and absence of a target polynucleotide/polynucleotide hybrid are combined to produce a reaction mixture. The rate of change in an optical property of the dye in the reaction mixture is compared to a reference value characteristic of the rate of change in the optical property of the dye in a similar reaction mixture containing a known amount of a polynucleotide/polynucleotide hybrid to determine a relative rate of change in the optical property. The relative rate of change in the optical property of dye in the reaction mixture is correlated with the presence or amount of the specified target polynucleotide in the sample.