Abstract
Time-resolved microspectrofluorimetry and fluorescence microscopy imaging—two complementary fluorescence techniques—provide important information about the intracellular distribution, level of uptake and binding/interactions inside living cell of the labeled molecule of interest. They were employed to monitor the “fate” of AS1411 aptamer labeled by ATTO 425 in human living cells. Confocal microspectrofluorimeter adapted for time-resolved intracellular fluorescence measurements by using a phase-modulation principle with homodyne data acquisition was employed to obtain emission spectra and to determine fluorescence lifetimes in U-87 MG tumor brain cells and Hs68 non–tumor foreskin cells. Acquired spectra from both the intracellular space and the reference solutions were treated to observe the aptamer localization and its interaction with biological structures inside the living cell. The emission spectra and the maximum emission wavelengths coming from the cells are practically identical, however significant lifetime lengthening was observed for tumor cell line in comparison to non–tumor one.
Similar content being viewed by others
References
Goodchild J (2011) Therapeutic Oligonucleotides. In: Goodchild J (ed) Methods in Molecular Biology, Volume 764, Springer Protocols, Humana Press, pp 1–15
Nimjee SM, Rusconi CP, Sullenger BA (2005) Aptamers: an emerging class of therapeutics. Annu Rev Med 56:555–583. doi:10.1146/annurev.med.56.062904.144915
Ashrafuzzaman M (2014) Aptamers as both drugs and drug-carriers. BioMed Res Int 2014:697923. doi:10.1155/2014/697923
Bates PJ, Laber DA, Miller DM, Thomas SD, Trent JO (2009) Discovery and development of the G-rich oligonucleotide AS1411 as a novel treatment for cancer. Exp Mol Pathol 86:151–164. doi:10.1016/j.yexmp.2009.01.004
Reyes-Reyes EM, Teng Y, Bates PJ (2010) A new paradigm for aptamer therapeutic AS1411 action: uptake by macropinocytosis and its stimulation by a nucleolin-dependent mechanism. Cancer Res 70:8617–8629. doi:10.1158/0008-5472.CAN-10-0920
Praus P, Sureau F (2000) Spectral decomposition of intracellular complex fluorescent signals using multiwavelength phase modulation lifetime determination. J Fluor 10:361–364
Tran PLT, De Cian A, Gros J, Moriyama R, Mergny JL (2013) Tetramolecular quadruplex stability and assembly in Quandruplex Nucleic Acids. In: Chaires JB and Graves D (ed), Topics in Current Chemistry 330, Springer-Verlag Berlin Heidelberg, pp 243–274. doi: 10.1007/128_2012_334
Kočišová E, Antalík A, Procházka M (2013) Drop coating deposition Raman spectroscopy of liposomes: role of cholesterol. Chem Phys Lipids 172–173: 1–5. doi: 10.1016/j.chemphyslip.2013.04.002
Praus P, Kočišová E, Seksek O, Sureau F, Štěpánek J, Turpin PY (2007) Advanced microfluorescence methods in monitoring intracellular uptake of “antisense” oligonucleotides. Curr Org Chem 11:515–527. doi:10.2174/138527207780368210
Lakowicz JR (1999) Principles of fluorescence spectroscopy. Kluwer Academic/Plenum Publishers, New York
Kočišová E, Praus P, Rosenberg I, Seksek O, Sureau F, Štěpánek J, Turpin PY (2004) Intracellular uptake of modified oligonucleotide studied by two fluorescence techniques. Biopolymers 74:110–114. doi:10.1002/bip.20055
Acknowledgments
Financial support from the Grant Agency of the Czech Republic project No. 15-06785S and from the Czech Ministry of Education Youth and Sports project No. 7AMB14FR020 is gratefully acknowledged.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Kočišová, E., Praus, P., Bok, J. et al. Intracellular Monitoring of AS1411 Aptamer by Time-Resolved Microspectrofluorimetry and Fluorescence Imaging. J Fluoresc 25, 1245–1250 (2015). https://doi.org/10.1007/s10895-015-1612-3
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10895-015-1612-3