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Anal Chem. 2012 Nov 6;84(21):9222-9. doi: 10.1021/ac3017423. Epub 2012 Oct 22.

Spectrally resolved chemiluminescent probes for sensitive multiplex molecular quantification.

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Gen-Probe Incorporated, 10210 Genetic Center Drive, San Diego, California 92121, United States.


Luminophores are frequently utilized probe labels for detecting biological analytes. Multiple fluorescent luminophores, or fluorophores, can be readily distinguished from one another based on different energy excitation and emission wavelengths and lifetimes. However, suitable methods and reagents for distinguishing multiples of the much more sensitive chemically initiated luminophores have been limited. Herein we describe a new class of hybrid luminophore probes that emit light of distinct wavelength ranges and intensities upon energy transfer (ET) from an in-common, acridinium ester chemiluminophore to a covalently conjugated fluorophore. This format supports rapid, rational design of spectrally resolvable, chemically initiated probes. Time-resolved spectrographic and luminescence characterizations indicate that ET is not dependent on overlap in the emission spectrum of the luminophore and the absorption spectra of acceptors, suggesting a non-Förster resonance ET mechanism. Analysis of a combination of the chemiluminophore and new chemiluminophore-acceptor conjugate probes demonstrates the benefits of their use in sensitive, multiplex quantification of nucleic acid sequences indicative of environmentally relevant microbes without prior enzymatic amplification.

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