Hole-burning and picosecond time-resolved spectroscopy of isolated molecular clusters

Stacey A. Wittmeyer; Andrew J. Kaziska; Maria I. Shchuka; Michael R. Topp

doi:10.1117/12.44250

1 July 1991 Hole-burning and picosecond time-resolved spectroscopy of isolated molecular clusters

Stacey A. Wittmeyer, Andrew J. Kaziska, Maria I. Shchuka, Michael R. Topp

Proceedings Volume 1435, Optical Methods for Ultrasensitive Detection and Analysis: Techniques and Applications; (1991) https://doi.org/10.1117/12.44250
Event: Optics, Electro-Optics, and Laser Applications in Science and Engineering, 1991, Los Angeles, CA, United States

Abstract

A precolumn derivatization method has been developed for high performance liquid chromatographic (HPLC) analysis of DNA damage using fluorescence detection. The modified nucleotide, having excised enzymatically from the exposed DNA, is enriched from the normal nucleotides and labeled with a fluorescent reagent. The labeling procedure involves phosphoramidation of the nucleotide with ethylenediamine (EDA) followed by conjugation of the free amino end of the phosphoramidate with 5-dimethylaminonaphthalene 1-sulfonyl chloride, commonly known as Dansyl chloride. The dansylated nucleotide can be analyzed with a sub-picomole limit of detection (LOD) by conventional HPLC using a conventional fluorescence detector. By combining microbore HPLC with laser-induced fluorescence (LIF) detection, we present the development of an analytical system that has sub-femtomole LOD for real-time analysis of the dansylated nucleotide. The application of the developed system in fluorescence postlabeling assay of a small alkyl-modified nucleotide (5-methyldCMP) in calf-thymus DNA is discussed.

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Stacey A. Wittmeyer, Andrew J. Kaziska, Maria I. Shchuka, and Michael R. Topp "Hole-burning and picosecond time-resolved spectroscopy of isolated molecular clusters", Proc. SPIE 1435, Optical Methods for Ultrasensitive Detection and Analysis: Techniques and Applications, (1 July 1991); https://doi.org/10.1117/12.44250

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KEYWORDS

Hole burning spectroscopy

Luminescence

Molecules

Picosecond phenomena

Methane

Molecular spectroscopy

Spectroscopy

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