Danying Lin

Quantitative Analysis of Intracellular Calcium and Mitochondrial Kinetic Fluorescence Changes in GSNO-induced Thymocyte Early Apoptosis

A fluorescence microscopy imaging technique was applied to observe the single-cell kinetic changes of intracellular Ca2+ concentration ([Ca2+]i) and mitochondrial membrane potential (ΔΨm) during the early stage of S-nitrosoglutathione (GSNO)-induced thymocytes apoptosis. The kinetic features of [Ca2+]i and ΔΨm were quantitatively analyzed and compared by fitting the fluorescence intensity data. The mathematical parameter, inflection point which indicated the time point when [Ca2+]i or ΔΨm changed the most rapidly, was proposed to analyze the fitting curve. The results revealed that the inflection point of [Ca2+]i always appeared prior to that of ΔΨm during apoptosis induced by a certain GSNO concentration. Both the [Ca2+]i and ΔΨm changed in a GSNO concentration-dependent manner. Another parameter, half-max effect point was also employed and displayed the similar results. Such quantitative analyses of real-time observations at the single-cell level are useful for interpreting the sequence of the biological events operating in GSNO-induced thymocyte apoptosis.

Three-Dimensional Imaging and Tracking for Constrained Brownian Motion of Single Nano-Scaled Particles in Solution

Objective-type total internal reflection fluorescence technique and Gaussian fit single particle tracking algorithm have been used to study the constrained Brownian motion characteristics of single fluorescent microspheres of 40 nm diameter in real-time. Three-dimensional tracking has been realized from the two-dimensional images by utilizing the exponential decay of the fluorescence intensity excited by the evanescent field. Trajectory analyses have shown that single fluorescent beads diffused in a constrained Brownian mode with a diffusion coefficient a little smaller than theoretical estimates in the plane-parallel direction.

High Sensitivity Real‐Time Fluorescence Microscopy and its Application in Thymocyte Apoptosis (abstract)

Real‐time observation of molecules in living cells is of great importance in the field of biomedicine. Fluorescent microscopic imaging techniques are highly sensitive and specific, and allow detailed visualization. However, applicable methods still face challenges due to cellular complexity. Our interests include working to meet these challenges to study real‐time and in situ imaging of special biomolecules in living cells. Specific applications of fluorescent microscopic imaging have been explored, such as thymocyte apoptosis induced by S‐nitrosoglutathione. Start‐up processes of thymocyte apoptosis and apoptosis‐initiating time points were recorded in real‐time at the single living cell level. Real‐time studies of the cooperation processes of the inducer with various inhibitors suggest different mechanisms of action. During the apoptosis‐inducing course, rapid increase of intracellular free calcium ion concentration and gradual loss of mitochondrial membrane potential were also detected in real‐time and...

Real-time Observation of Intracellular Calcium Changes in GSNO-induced Mouse Thymocyte Early Apoptosis

Using fast fluorescence micro-imaging technique, we have studied the real-time changes of intracellular Ca2+ concentration ([Ca2+]i) in S-nitrosoglutathione (GSNO)-induced mouse thymocyte early apoptosis. It was observed that [Ca2+]i increases rapidly until reaches a platform. The variety range and velocity are related to the GSNO concentration and the way how to treat with the thymocytes. These results show a great difference from the dexamethasone (DEX)-induced thymocytes early apoptosis, caused by the different apoptosis mechanisms. This article supplies a new method for investigating the mechanism in GSNO-induced mouse thymocyte apoptosis.

Two-dimensional Imaging and Tracking of Single Quantum dot-FRET

Using a custom-made setup ICCD-DualView-TIRFM system, we have successfully performed two-dimensional imaging and tracking of fluorescence resonance energy transfer (FRET)-occurring single quantum dot and its energy acceptor Cy5. This work laid a foundation for the further application of quantum dot-FRET in life sciences.