Research Center for Analytical Sciences, Department of Chemistry, College of Sciences , Northeastern University , Shenyang 110819 , China.
Anal Chem. 2019 Mar 19;91(6):4157-4163. doi: 10.1021/acs.analchem.9b00018. Epub 2019 Feb 27.
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 Zn aptamer, whereas the receptor FAM is conjugated with the complementary DNA sequence to ensure selectivity. MCF-7 cells are cultured sequentially with Zn (20, 40, 55, 70, 85, and 100 μmol L) and CDs-FAM (100 μg mL), which is used for fluorescence imaging (at λ = 405 nm and λ = 440-490 nm for CDs, λ = 500-550 nm for FAM) to provide a relative fluorescence ratio (( F - F)/ F, F = I/ I), 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 Zn by MCF-7 cells is further evaluated with this approach by culturing with 100 μmol L of Zn 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.
细胞金属的荧光成像已有广泛报道。然而，利用荧光成像定量细胞内金属目前还不可行，极具挑战性。在这项工作中，基于碳点（CDs）到荧光素（FAM）的荧光共振能量转移（FRET），设计了一种带有两个荧光标记互补 DNA 链的比率探针，用于通过荧光成像定量细胞内锌。供体 CDs 用锌适配体修饰，而受体 FAM 则与互补 DNA 序列缀合，以确保选择性。将 MCF-7 细胞依次用 Zn（20、40、55、70、85 和 100 μmol L）和 CDs-FAM（100 μg mL）培养，用于荧光成像（λ=405nm 和 λ=440-490nm 用于 CDs，λ=500-550nm 用于 FAM），以提供相对荧光比（( F - F)/ F，F = I/ I），随后用 ICPMS 定量细胞内锌。荧光图像中的相对荧光比与 ICPMS 得出的细胞内锌含量之间呈现线性相关，这便于通过荧光成像进行细胞内锌定量。它在细胞培养过程或体内特别有助于实时追踪细胞内锌。通过用 100 μmol L 的 Zn 培养不同时间来进一步评估 MCF-7 细胞对 Zn 的摄取，在孵育 60 分钟时观察到每个细胞的最大摄取量为 60.5 fg。这一值也通过 ICPMS 检测得到了进一步证实。
