Yuqiang Jiang

Recent Advances in Super-Resolution Fluorescence Imaging and Its Applications in Biology

Fluorescence microscopy has become an essential tool for biological research because it can be minimally invasive, acquire data rapidly, and target molecules of interest with specific labeling strategies. However, the diffraction-limited spatial resolution, which is classically limited to about 200 nm in the lateral direction and about 500 nm in the axial direction, hampers its application to identify delicate details of subcellular structure. Extensive efforts have been made to break diffraction limit for obtaining high-resolution imaging of a biological specimen. Various methods capable of obtaining super-resolution images with a resolution of tens of nanometers are currently available. These super-resolution techniques can be generally divided into three primary classes: (1) patterned illumination-based super-resolution imaging, which employs spatially and temporally modulated illumination light to reconstruct sub-diffraction structures; (2) single-molecule localization-based super-resolution imaging, which localizes the profile center of each individual fluorophore at subdiffraction precision; (3) bleaching/blinking-based super-resolution imaging. These super-resolution techniques have been utilized in different biological fields and provide novel insights into several new aspects of life science. Given unique technical merits and commercial availability of super-resolution fluorescence microscope, increasing applications of this powerful technique in life science can be expected.

Trapping and manipulation of a single micro-object in solution with femtosecond laser-induced mechanical force

A nondestructive and noncontact method to trap and manipulate a microparticle in solution is proposed by utilizing femtosecond laser-induced nonlinear phenomena. A 90μm diameter polystyrene bead in solution was trapped and manipulated by scanning femtosecond laser pulses around it, which was ascribed to shockwave, cavitation bubble, and jet flow. The maximum mechanical force exerted by laser irradiation was estimated to be over 1μN. In comparison with conventional optical tweezers, this method provides not only a much larger trapping force but also a noninvasive advantage.

Intracellular Temperature Sensing: An Ultra-bright Luminescent Nanothermometer with Non-sensitivity to pH and Ionic Strength.

Luminescence thermometry usually suffer from cellular complexity of the biochemical environment (such as pH and ionic strength), and thus the accuracy and reliability of the determined intracellular temperature are directly affected. Herein, a photoluminescent nanothermometer composed of polymer encapsulated quantum dots (P-QD) has been developed. And the prepared nanothermometer exhibits some advantages: such as non-sensitivity to pH and ionic strength, as well as high detection sensitivity and ultrahigh reversibility. The intracellular temperature was accurately determined under physiological conditions with different pH and ionic strength, and direct measurement of thermogenesis in individual cells has been achieved.

Extremely High Brightness from Polymer-Encapsulated Quantum Dots for Two-photon Cellular and Deep-tissue Imaging

Materials possessing high two photon absorption (TPA) are highly desirable for a range of fields, such as three-dimensional data storage, TP microscopy (TPM) and photodynamic therapy (PDT). Specifically, for TPM, high TP excitation (TPE) brightness (σ × ϕ, where σ is TPA cross-sections and ϕ is fluorescence quantum yield), excellent photostability and minimal cytotoxicity are highly desirable. However, when TPA materials are transferred to aqueous media through molecule engineering or nanoparticle formulation, they usually suffer from the severely decrease of quantum yield (QY). Here, we report a convenient and efficient method for preparing polymer-encapsulated quantum dots (P-QD). Interestingly, the QY was considerably enhanced from original 0.33 (QDs in THF) to 0.84 (P-QD in water). This dramatic enhancement in QY is mainly from the efficiently blocking nonradiative decay pathway from the surface trap states, according to the fluorescence decay lifetimes analysis. The P-QD exhibits extremely high brightness (σ × ϕ up to 6.2 × 106 GM), high photostability, excellent colloidal stability and minimal cytotoxicity. High quality cellular TP imaging with high signal-to-background ratio (> 100) and tissue imaging with a penetration depth of 2200 μm have been achieved with P-QD as probe.

Two-Photon Luminescence and Second Harmonic Generation from Gold Micro-Plates

Micron-sized gold plates were prepared by reducing chloroauric acid with lemongrass extract. Their two-photon luminescence (TPL) and second harmonic generation (SHG) were investigated. The results show that the TPL and SHG intensity of gold plates is dependent on the wavelength and polarization of excitation laser. The TPL intensity of gold plates decreases with the increase of the excitation wavelength except for a small peak around 820–840 nm, while SHG intensity increases with the excitation wavelength redshift. In addition, it is found that the TPL intensity of the gold plate’s edge is related with the angle between the edge orientation and the polarization direction of the excitation light. The TPL intensity increases with the angle increase from 0° to 90°.

CAMSAP3-dependent microtubule dynamics regulates Golgi assembly in epithelial cells

The Golgi assembly pattern varies among cell types. In fibroblast cells, the Golgi apparatus concentrates around the centrosome that radiates microtubules; whereas in epithelial cells, whose microtubules are mainly noncentrosomal, the Golgi apparatus accumulates around the nucleus independently of centrosome. Little is known about the mechanisms behind such cell type-specific Golgi and microtubule organization. Here, we show that the microtubule minus-end binding protein Nezha/CAMSAP3 (calmodulin-regulated spectrin-associated protein 3) plays a role in translocation of Golgi vesicles in epithelial cells. This function of CAMSAP3 is supported by CG-NAP (centrosome and Golgi localized PKN-associated protein) through their binding. Depletion of either one of these proteins similarly induces fragmentation of Golgi membranes. Furthermore, we find that stathmin-dependent microtubule dynamics is graded along the radial axis of cells with highest activity at the perinuclear region, and inhibition of this gradient disrupts perinuclear distribution of the Golgi apparatus. We propose that the assembly of the Golgi apparatus in epithelial cells is induced by a multi-step process, which includes CAMSAP3-dependent Golgi vesicle clustering and graded microtubule dynamics.

Proposal of a wavelength filter with a cut corner based on Equilateral-Triangle-Resonator.

We propose an equilateral triangle resonator filter with an output waveguide and analyzed by the finite-difference time-domain technique. The filter can realize directional output with a high Q mode by means of the mode-field coupled into the output waveguide, which results a reduction in the scattering loss at the vertices. In addition, to the deformed equilateral triangle resonator filter, an optimum parameter with a cut corner of 0.23 μm, which is equal to that of the input waveguide and can be an optimal cut, is found to help increase in finesse, Q factors, extinction ratio and the output intensity on resonance of the drop port normalized with the through port .

SECONDARY CONVERGENCE IN FEMTOSECOND LASER TRAPPING

When femtosecond laser pulses pass through a trapped polystyrene bead, water breakdown is induced even though the energy of laser pulse is much lower compared to the threshold value of breakdown when the femtosecond laser directly irradiates in water. This mechanism is assigned to the secondary convergence of the laser by the trapped bead.

Femtosecond Laser Manipulation Techniques for Individual Patterning of Biological Micro-object

Several kinds of manipulation of biological cells were performed utilizing regeneratively amplified femtosecond laser system. When single-shot pulse of an amplified Ti: Sapphire femtosecond laser pulse is focused on a culture medium, shockwave and cavitation bubble are generated with little heating. An impulsive force resulting in these phenomena was applied to pttern specific cells form a culture substrate. Furthermore, laser trapping of cells was realized using high-repetition rate pulses from the laser oscillator. Although the cell was trapped stably when the laser power was less than 100 mW, the cell was burst above the threshold laser power. The bursting would be due to heating inside cell, on which the laser was focused and multiphoton absorption was induced. On the bases of these results, we propose a new methodology to pattern biological cells, which is speedy and flexible when compared with previous micropatterning methods.

Effects of nonlinear polarization in optical trapping

Conventional optical tweezers handle non-absorptive (transparent) particles, and the force exerted to the particle is proportional to the intensity of the trapping laser. Recently, some works on selective trapping/manipulation of absorptive particles based on resonance effects have been reported. Some years ago, our research group found that the use of an ultra-short pulsed laser as a trapping light source shows nonlinear optical effects on trapping behavior for gold nanoparticles under pre-resonant conditions [1]. In this paper, we briefly describe the experimental observation of the phenomenon and interpretation of that.