Two-photon fluorescence cross-correlation spectroscopy as a potential tool for high-throughput screening of DNA repair activity

Maddalena Collini; Michele Caccia; Giuseppe Chirico; Flavia Barone; Eugenia Dogliotti; Filomena Mazzei

doi:10.1093/nar/gni166

Two-photon fluorescence cross-correlation spectroscopy as a potential tool for high-throughput screening of DNA repair activity

Maddalena Collini, Michele Caccia, Giuseppe Chirico, Flavia Barone, Eugenia Dogliotti, Filomena Mazzei

Istituto Europeo di Oncologia

Research output: Contribution to journal › Article › peer-review

Abstract

Several lines of evidence indicate that differences in DNA repair capacity are an important source of variability in cancer risk. However, traditional assays for measurement of DNA repair activity in human samples are laborious and time-consuming. DNA glycosylases are the first step in base excision repair of a variety of modified DNA bases. Here, we describe the development of a new sensitive DNA glycosylase assay based on fluorescence cross-correlation spectroscopy (FCCS) with two-photon excitation. FCCS was applied to the measurement of uracil DNA glycosylase activity of human cell extracts and validated by comparison with standard gel electrophoresis assay. Our results indicate that FCCS can be adapted to efficient assays for DNA glycosylase activity in protein extracts from human cells. This method has a potential for the development of automated screening of large number of samples.

Original language	English
Pages (from-to)	1-8
Number of pages	8
Journal	Nucleic Acids Research
Volume	33
Issue number	19
DOIs	https://doi.org/10.1093/nar/gni166
Publication status	Published - 2005

ASJC Scopus subject areas

Genetics

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abstract = "Several lines of evidence indicate that differences in DNA repair capacity are an important source of variability in cancer risk. However, traditional assays for measurement of DNA repair activity in human samples are laborious and time-consuming. DNA glycosylases are the first step in base excision repair of a variety of modified DNA bases. Here, we describe the development of a new sensitive DNA glycosylase assay based on fluorescence cross-correlation spectroscopy (FCCS) with two-photon excitation. FCCS was applied to the measurement of uracil DNA glycosylase activity of human cell extracts and validated by comparison with standard gel electrophoresis assay. Our results indicate that FCCS can be adapted to efficient assays for DNA glycosylase activity in protein extracts from human cells. This method has a potential for the development of automated screening of large number of samples.",

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AU - Chirico, Giuseppe

AU - Barone, Flavia

AU - Dogliotti, Eugenia

AU - Mazzei, Filomena

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Two-photon fluorescence cross-correlation spectroscopy as a potential tool for high-throughput screening of DNA repair activity

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