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The effects of temperature, salt concentration, and formamide concentration on the emission characteristics of commonly used fluorescent labels were evaluated on DNA microchips. The emission intensities of different fluorophores without hybridization were observed to vary, each to a different extent, to mainly temperature changes. Rhodamine red, TAMRA (tetramethylrhodamine), and dyes from the carbocyanide group exhibited the largest variations, and Texas Red and Oregon Green exhibited the smallest variations. This temperature dependency was shown to affect results obtained during melting curve analysis in DNA microarray studies. To minimize the bias associated with the temperature-dependent emission of different fluorescent labels, a normalization step was proposed.

Emission Characteristics of Fluorescent Labels with Respect to Temperature Changes and Subsequent Effects on DNA Microchip Studies