| Dedication | 第4-5页 |
| Acknowledgements | 第5-6页 |
| 摘要 | 第6-10页 |
| ABSTRACT | 第10-15页 |
| Chapter 1. Introduction of fluorescence bioimaging techniques and fluorescent probes | 第20-52页 |
| 1.1. Bioimaging/ Fluorescence bioimaging | 第20页 |
| 1.2. Fluorescence Microscopy | 第20-27页 |
| 1.2.1. Wide-field fluorescence microscopy (WFFM) | 第21-22页 |
| 1.2.2. Confocal microscopy | 第22-23页 |
| 1.2.3. Two-photon fluorescence microscopy | 第23-27页 |
| 1.2.3.1. Two-photon absorption/excitation | 第23-25页 |
| 1.2.3.2. Two-photon microscopy instrumentation | 第25-26页 |
| 1.2.3.3. Applications of TPM | 第26-27页 |
| 1.3. Fluorescence-lifetime imaging microscopy (FLIM) | 第27-30页 |
| 1.3.1. FRET Imaging | 第28-29页 |
| 1.3.2. Time-Correlated Single Photon Counting (TCSPC) | 第29-30页 |
| 1.4. Super-resolution microscopy (SRM) | 第30-32页 |
| 1.4.1. Stimulated emission depletion (STED) microscopy | 第31-32页 |
| 1.5. Fluorescent bioimaging probes | 第32-39页 |
| 1.5.1. Small organic two-photon fluorescence probe | 第33-34页 |
| 1.5.2. Two-photon fluorescence probes for subcellular targeting | 第34-36页 |
| 1.5.3. Two-photon fluorescent probes to detect intracellular metal ions | 第36页 |
| 1.5.4. Two-photon fluorescent probes for nucleic acids (NAs) staining | 第36-37页 |
| 1.5.5. Detection of microenvironment in cells by two-photon fluorescence probes | 第37-38页 |
| 1.5.6. Two-photon metal complexes | 第38-39页 |
| 1.6. Aims and Outlines of thesis | 第39-43页 |
| References | 第43-52页 |
| Chapter 2. A series of water-soluble pyridinium derivatives with two-photon absorption in the nearinfrared region for mitochondria targeting under stimulated emission depletion (STED) nanoscopy | 第52-88页 |
| 2.1. Introduction | 第52-54页 |
| 2.2. Experimental Methods | 第54-59页 |
| 2.2.1. Materials and apparatus | 第54页 |
| 2.2.2. Synthetic procedures of NL1-3 and PL1-3 | 第54-57页 |
| 2.2.3. Cell Culture | 第57页 |
| 2.2.4. Cytotoxicity and photostability of NL1-3 and PL1-3 | 第57-58页 |
| 2.2.5. Animal studies | 第58页 |
| 2.2.6. Cell imaging using confocal laser scanning microscopy | 第58页 |
| 2.2.7. Stimulated emission depletion (STED) nanoscopy | 第58-59页 |
| 2.3. Results and discussions | 第59-82页 |
| 2.3.1. Crystal structures, UV-Vis absorption and emission spectra, and TD-DFT studies of the targetcompounds | 第59-67页 |
| 2.3.2. One-photon emission and two-photon absorption cross section response to solvent viscosity | 第67-74页 |
| 2.3.3. Biological imaging applications of NL1 | 第74-82页 |
| 2.3.3.1. Costaining with Mito Tracker Red and consistency with mitochondrial membrane potential | 第77-80页 |
| 2.3.3.2. Application of NL1 on STED nanoscopy | 第80-82页 |
| References | 第82-88页 |
| Chapter 3. A Series of two-photon absorption pyridinium sulfonate inner salts targetingendoplasmic reticulum (ER), inducing cellular stress and mitochondria-mediated apoptosis incancer cells | 第88-115页 |
| 3.1. Introduction | 第88-90页 |
| 3.2. Experimental | 第90-95页 |
| 3.2.1. Materials and apparatus | 第90页 |
| 3.2.2. Synthetic procedures of TriphenER1-2 and DiphenthioER1-2 | 第90-92页 |
| 3.2.3. Fluorescence-activated cell sorter (FACS) analysis of cellular uptake | 第92页 |
| 3.2.4. Cell viability/proliferation assay | 第92-93页 |
| 3.2.5. Cell Culture | 第93页 |
| 3.2.6. Cell imaging using confocal laser scanning microscopy | 第93页 |
| 3.2.7. Stimulated emission depletion (STED) nanoscopy | 第93-94页 |
| 3.2.8. Western blot | 第94-95页 |
| 3.3. Results and discussion | 第95-111页 |
| 3.3.1. Crystal structures | 第95-97页 |
| 3.3.2. Photophysical properties | 第97-100页 |
| 3.3.3. Lipophilicity and cellular uptake of TriphenER1-2 and DiphenthioER1-2 | 第100-103页 |
| 3.3.4. Intracellular localization of DiphenthioER1 and two-photon imaging application | 第103-105页 |
| 3.3.5. DiphenthioER1 induced endoplasmic reticulum stress and nuclear misshaping | 第105-108页 |
| 3.3.6. Mitochondrial fragmentation and apoptosis | 第108-111页 |
| References | 第111-115页 |
| Chapter 4. DiphenthioER1;a two-photon active pyridinium derivative causing opening of bloodbrain barriers via junctional proteins downregulation | 第115-130页 |
| 4.1. Introduction | 第115-116页 |
| 4.2. Experimental Section | 第116-121页 |
| 4.2.1. Cell culture and subculture | 第116-117页 |
| 4.2.2. Cytotoxicity of DiphenthioERl towards bEND.3 cells | 第117页 |
| 4.2.3. Immunofluorescence (IF) | 第117页 |
| 4.2.4. 3D in vitro BBB model set up | 第117-119页 |
| 4.2.5. Preparation of trans-well slides for microscopy: Protocol | 第119-121页 |
| 4.3. Results and Discussions | 第121-127页 |
| 4.3.1. Cytotoxicity and cellular uptake of DiphenthioER1 | 第121-122页 |
| 4.3.2. Effect of DiphenthioERl on tight junction (TJ) protein expression in 2D and 3D cultures | 第122-127页 |
| 4.3.2.1. Immunofluorescence labelling for claudin-5 in 2D culture and 3D cultures | 第123-125页 |
| 4.3.2.2. Immunofluorescence labelling for zo-1 in 2D culture | 第125-127页 |
| References | 第127-130页 |
| Chapter 5. SL-Neu; a Two-photon Fluorescent Chemical Probe for NeuN Specific Live Neuron Labeling | 第130-151页 |
| 5.1. Introduction | 第130-131页 |
| 5.2. Experimental | 第131-134页 |
| 5.2.1. Reagents and instruments | 第131-132页 |
| 5.2.2. Synthesis and characterization | 第132-133页 |
| 5.2.3. Animal studies | 第133页 |
| 5.2.4. Cell imaging using confocal laser scanning microscopy | 第133-134页 |
| 5.2.5. Stimulated emission depletion (STED) nanoscopy | 第134页 |
| 5.3. Results and discussions | 第134-149页 |
| 5.3.1. Crystal structure and analysis | 第134-138页 |
| 5.3.2. Linear optical properties | 第138-142页 |
| 5.3.2.1. UV-visible spectrum | 第138-139页 |
| 5.3.2.2. Density functional theory calculations | 第139-140页 |
| 5.3.2.3. One photon fluorescence spectroscopy | 第140-142页 |
| 5.3.3. Nonlinear optical properties | 第142-143页 |
| 5.3.3.1. Two-photon induced fluorescence spectroscopy and its verification | 第142-143页 |
| 5.3.3.2. Two-photon absorption cross section | 第143页 |
| 5.3.4. Biological applications of SL-Neu | 第143-149页 |
| 5.3.4.1. SL-Neu uptake by different brain regions | 第143-144页 |
| 5.3.4.2. Colocalization of SL-Neu with DAPI and β-Ⅲ-tubulin | 第144-146页 |
| 5.3.4.3. Involvement of non-neuronal and neuronal cells | 第146-148页 |
| 5.3.4.4. Confirmation of SL-Neu specific binding under STED | 第148-149页 |
| References | 第149-151页 |
| Chapter 6: Characterization and biological applications of Terpyridine Mn~(2+)Complexes | 第151-171页 |
| 6.1. Introduction | 第151页 |
| 6.2. Experimental section | 第151-156页 |
| 6.2.1. Reagents and instruments | 第151-152页 |
| 6.2.2. Synthesis of mononuclear Mn complexes | 第152-154页 |
| 6.2.3. Determination of size and morphology of Mn1-Mn4 | 第154页 |
| 6.2.4. Cytotoxicity by MTT assay | 第154-155页 |
| 6.2.5. Cell culture | 第155页 |
| 6.2.6. Cell imaging using confocal laser scanning microscopy | 第155-156页 |
| 6.3. Results and discussion | 第156-169页 |
| 6.3.0. Single crystal crystal structure and structure analysis | 第156页 |
| 6.3.1. UV-vis absorption spectra of terpyridine-manganese complexes | 第156-159页 |
| 6.3.2. One photon fluorescence spectroscopy | 第159-161页 |
| 6.3.3. Fluorescence quantum yield and fluorescence lifetime | 第161-162页 |
| 6.3.4. Two-photon induced fluorescence | 第162-164页 |
| 6.3.5. Size and morphology determination by DLS, SEM and TEM | 第164-167页 |
| 6.3.6. Cytotoxicity Assay | 第167页 |
| 6.3.7. Cellular imaging application of Mn4 | 第167-169页 |
| References | 第169-171页 |
| Conclusions | 第171-174页 |
| Publications | 第174页 |
