| The previous study found that a small chemical or physical change within the cells may indicate a significant change in the life of the body. So with the rapid development of the life science, a precise, sensitive and rapid detection of the small molecules, ions, nucleic acids or proteins in single cell or even organelle are more and more urgent. Fluorescence sensing technology is a hot research of biological analysis at present, for its advantages of high selectivity, high sensitivity, fast in situ real-time analysis and no damage to the sample. It has been proved that some new fond fluorescent molecular probes have highly sensitive and special detection of molecules, ions and so on, which provides a further technical support for the development of the biological analysis technology.In this paper, by the research of fluorescent probes, we firstly designed a pH probe FR-Lys based on Oxygen derivatives, which realized the location of lysosomal and detection of the dynamic changes of pH in it. Secondly we designed two ratiometric two-photon probes Np-Rh and Np-Rh-Pd based on Through-bond energy transfer (TBET), for special detection and deep-tissue imaging of Cu2+ions and Pd2+ ions respectively. The main contents are as follows:1. In Chapter 2, we developed a lysosomal-targeting pH fluorescent probe (FR-Lys) through a smart integration of the xanthene, benzothiazole and morpholine. In this probe, morpholine group serves as a targeting unit for lysosome; the xanthane derivative exhibits a pH-modulated open/close reaction of the spirocycle, while the o-hydroxy benzoxazole moiety shows a pH modulated excited-state intramolecular proton transfer (ESIPT) process. Such a design affords the probe a ratiometric fluorescence response towards pH with pH values ranging from 4.0 to 6.3. The response process of FR-Lys to pH is fast and reversible, Moreover the probe has further advantages of easy synthesis, high photostability and low cytotoxicity, which qualify it to apply for dynamic imaging of pH changes in lysosomes with satisfactory results.2. In Chapter 3, we connect the two-photon fluorophore naphthalene derivatives and Rhodamine B derivative with a structure of spirolactam ring, for building a ratiometric two-photon platform with a structure of D-?-A. There is an efficient energy transfer by Through-bond energy transfer on the platform, high enough to 90%. In the meantime, we get two well-resolved emission peaks separated by 100 nm and large two-photon active absorption cross-section. We use common Cu2+ ions as the objects of detection to construct a fluorescent probe Np-Rh. Experimental results show that the probe has specific detection of the Cu2+ ions, the detection limit is 3.0 x 10-7 M. The stability of Np-Rh under the physiological conditions is good, and the probe realized two-photon imaging of living cells and tissues and showed high ratiometric imaging resolution and deep-tissue imaging depth of 70-180 ?m.3. In Chapter 4, we design a ratiometric two-photon probe for Pd2+ ions (Np-Rh-Pd) based on the platform in the third part. When Pd2+ ions exist, the N, O atoms on the donor (rhodamine B spirolactams) will coordinate with them, which makes the emission peak of the probe red shift 100 nm. The signal-to-background ratio of the detection to Pd2+ ions was 31.2, with a detection limit of 2.3×10-7 M. The probe realized two-photon imaging of living cells and tissues and showed high ratiometric imaging resolution and deep-tissue imaging depth of 90-270 ?m... |
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