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BioMed Research International
Volume 2014, Article ID 196837, 10 pages
http://dx.doi.org/10.1155/2014/196837
Research Article

Reliable Assessment and Quantification of the Fluorescence-Labeled Antisense Oligonucleotides In Vivo

Department of Biomedical Engineering, National Cerebral and Cardiovascular Center Research Institute, Suita, Osaka 565-8565, Japan

Received 25 February 2014; Revised 22 April 2014; Accepted 22 April 2014; Published 25 May 2014

Academic Editor: Pradeep Kumar

Copyright © 2014 Maria Chiara Munisso and Tetsuji Yamaoka. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

The availability of fluorescent dyes and the advances in the optical systems for in vivo imaging have stimulated an increasing interest in developing new methodologies to study and quantify the biodistribution of labeled agents. However, despite these great achievements, we are facing significant challenges in determining if the observed fluorescence does correspond to the quantity of the dye in the tissues. In fact, although the far-red and near-infrared lights can propagate through several centimetres of tissue, they diffuse within a few millimetres as consequence of the elastic scattering of photons. In addition, when dye-labeled oligonucleotides form stable complex with cationic carriers, a large change in the fluorescence intensity of the dye is observed. Therefore, the measured fluorescence intensity is altered by the tissue heterogeneity and by the fluctuation of dye intensity. Hence, in this study a quantification strategy for fluorescence-labeled oligonucleotides was developed to solve these disadvantageous effects. Our results proved that upon efficient homogenization and dilution with chaotropic agents, such as guanidinium thiocyanate, it is possible to achieve a complete fluorescence intensity recovery. Furthermore, we demonstrated that this method has the advantage of good sensitivity and reproducibility, as well as easy handling of the tissue samples.