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Intracellular Zinc Quantification by Fluorescence Imaging with a FRET System.

Shu, Yang; Zheng, Na; Zheng, An-Qi; Guo, Ting-Ting; Yu, Yong-Liang; Wang, Jian-Hua.
Anal Chem; 91(6): 4157-4163, 2019 Mar 19.
Artigo em Inglês | MEDLINE | ID: mdl-30793593
Fluorescence imaging of cellular metals is widely reported. However, the quantification of intracellular metals with fluorescence imaging is so far not feasible and highly challenging. In this work, a ratiometric probe with two fluorescently labeled complementary DNA strains is designed for intracellular zinc quantification via fluorescence imaging, based on fluorescence resonance energy transfer (FRET) from carbon dots (CDs) to fluorescein (FAM). The donor CDs are modified with a Zn2+ aptamer, whereas the receptor FAM is conjugated with the complementary DNA sequence to ensure selectivity. MCF-7 cells are cultured sequentially with Zn2+ (20, 40, 55, 70, 85, and 100 µmol L-1) and CDs-FAM (100 µg mL-1), which is used for fluorescence imaging (at λex = 405 nm and λem = 440-490 nm for CDs, λem = 500-550 nm for FAM) to provide a relative fluorescence ratio (( F - F0)/ F0, F = ICDs/ IFAM), followed by quantifying intracellular zinc with ICPMS. A linear correlation is achieved between the relative fluorescence ratio in fluorescence images and the intracellular zinc content derived by ICPMS, which facilitates intracellular zinc quantification via fluorescence imaging. It is especially useful for real-time tracing of intracellular zinc during the cell culturing process or in vivo. The cellular uptake of Zn2+ by MCF-7 cells is further evaluated with this approach by culturing with 100 µmol L-1 of Zn2+ for different times, and a maximum uptake of 60.5 fg per cell is observed at an incubation time of 60 min. This value is further demonstrated well by ICPMS detection.
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