本论文介绍了一种基于扫描共聚焦显微镜和微流反应器用于研究化学振动子同步的系统。这个系统利用300微米的PNIPAm胶体颗粒作为振动子，Ru(vmbpy)(bpy)₂(PF₆)₂ 被参杂到振动子里充当BelousovZhabotinsky（BZ）振荡反应的催化剂。扫描共聚焦显微镜具有很高的灵敏度，可以给出高质量的图片以供研究分析。通过实验证明低于56W/cm²强度的扫描激光对BZ反应没有影响。这里所运用的微流反应器包括两个部分，PMMA材质的微池和PDMS材质的流道。此反应器可通过流道不停地补充BZ反应的反应物从而保证振荡的一致性。 / 通通过此系统，我们可以研究不同的两个振动子的同步问题。在实验中，振动子间的同步是由两者间的距离决定的。振动子在靠近时同步在分开至临界距离以外处不同步。另外，我利用COMSOL来模拟实验中的现象，发现模拟的结果和实验中的现象十分吻合。 / In this thesis, I present an experimental platform based on laser scanning confocal microscopy (LSCM) and continuous flow microreactor (CFMR) to study coupled chemical oscillators. PNIPAm gel particles around 300 micron were synthesized in the microfluidic device as the oscillators. Ru(vmbpy)(bpy)₂(PF₆)₂ was used as the catalyst of the BelousovZhabotinsky (BZ) reaction. The LSCM offers a good signal-to-noise ratio and better imaging quality. We demonstrated that the scanning laser with the power below 56W/cm² had no influence to BZ reaction. The CFMR, consisting of the PMMA microwell and the PDMS microchannel, can maintain the oscillation of the oscillators with a continuous supply of the BZ mixture. / The synchronization of the double heterogeneous oscillators was studied by the platform. The coupling intensity was controlled by changing the distance between the two oscillators. Results showed that the synchronization occurred as the oscillators were close and was lost as the oscillators were separated beyond a critical distance. The results of the numerical simulation by COMSOL agreed well with the experimental observation. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Guo, Dameng. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 40-41). / Abstracts also in Chinese. / Abstract --- p.i / 摘要 --- p.ii / Acknowledgement --- p.iii / Table of contents --- p.iv / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- Synchronization of chemical oscillating systems --- p.1 / Chapter 1.2 --- The Belousov-Zhabotinsky reaction --- p.4 / Chapter 1.2.1 --- FKN model --- p.5 / Chapter 1.2.2 --- Effect of light illumination on ruthenium catalyzed BZ reaction --- p.7 / Chapter 1.2.3 --- Luminescence of ruthenium catalyzed in the BZ reaction --- p.8 / Chapter 1.3 --- Oscillators based on the BZ reaction --- p.9 / Chapter 1.4 --- Detection methods for the BZ oscillating systems --- p.10 / Chapter 1.4.1 --- Ion selective electrode and optical microscopy --- p.10 / Chapter 1.4.2 --- Objective of the research --- p.10 / Chapter Chapter 2 --- LSCM and continuous flow microreactor based platform --- p.12 / Chapter 2.1 --- Introduction --- p.12 / Chapter 2.2 --- Experimental --- p.13 / Chapter 2.2.1 --- The fabrication of the oscillators --- p.13 / Chapter 2.2.2 --- The fabrication of the CFMR --- p.14 / Chapter 2.2.3 --- The detection method --- p.16 / Chapter 2.3 --- Results and discussions --- p.16 / Chapter 2.3.1 --- The comparison of PMMA and PDMS microreactors --- p.16 / Chapter 2.3.2 --- The flow rate of the BZ mixture to maintain the oscillation --- p.18 / Chapter 2.3.3 --- The size of the microreactor --- p.19 / Chapter 2.3.4 --- The factors to reduce the influence of the laser on the oscillators --- p.21 / Chapter 2.4 --- Conclusions --- p.23 / Chapter Chapter 3 --- The studying of the synchronization of the double oscillators --- p.24 / Chapter 3.1 --- Introduction --- p.24 / Chapter 3.1.1 --- Kuramoto model for illustrating the synchronization of double oscillators --- p.24 / Chapter 3.1.2 --- Transition from disorder to synchronization --- p.25 / Chapter 3.2 --- Experimental and simulation --- p.26 / Chapter 3.2.1 --- The dispensing and detection of the oscillators in the microreactor --- p.26 / Chapter 3.2.2 --- The simulation model --- p.27 / Chapter 3.3 --- Results and discussion --- p.28 / Chapter 3.3.1 --- The controlling of the coupling intensity --- p.28 / Chapter 3.3.2 --- The results of the synchronization --- p.30 / Chapter 3.3.2.1 --- Synchronization --- p.30 / Chapter 3.2.2.2 --- The critical distance for the synchronization and the transition --- p.31 / Chapter 3.3.3 --- The results of the simulation --- p.34 / Chapter 3.4 --- Conclusion --- p.35 / Chapter Chapter 4 --- Conclusion --- p.37 / Chapter 4.1 --- Summary --- p.37 / Chapter 4.2 --- Discussions and future perspectives --- p.38 / Reference --- p.40 / Appendix --- p.42 / Chapter 1. --- Code in Matlab for calculating the RGB value of ROI in the images from the LSCM --- p.42 / Chapter 2. --- The power of the laser --- p.43 / Chapter 2.1 --- The power of the laser in the LSCM --- p.43 / Chapter 2.2 --- The irradiation power on the oscillators --- p.44 / Chapter 3. --- The energy transferred to the oscillator --- p.44 / Chapter 4. --- The model in COMSOL --- p.45

Filamentous fungi, such as <i>Aspergillus nidulans</i>, are composed of tubular, highly polarized, multinucleate cells called hyphae. Polar growth involves secretion specifically at the hyphal tip. Secretion involves intracellular transport and co-ordination of the cytoskeleton and the endomembrane system. <p>Intracellular transport is likely mediated by cytoskeletal elements, which, in fungal cells consist primarily of actin and microtubules (MTs). An <i>A. nidulans</i> strain transformed with green fluorescent protein (GFP) tagged α-tubulin was utilized in the investigation of relationship between cytoplasmic MT arrays and hyphal growth rate. <i>A. nidulans</i> MTs were observed to be long and flexuous and to run roughly parallel to the long axis of hyphae. No correlation between relative MT abundance and hyphal growth rate was observed, although non-growing hyphae had a lower relative MT abundance than growing hyphae. Actin depolymerization decreased hyphal growth rate while MT depolymerization did not. MT stabilization increased hyphal growth rate. Ethanol, the solvent in which the MT and actin inhibitors were dissolved, increased both average overall growth rate and growth rate variability for individual hyphae. Taxol appeared to interact with irradiation to decreased growth rate during imaging. <p>Golgi are involved in secretion and potentially in polar growth. An <i>A. nidulans</i> α-coatomer protein (COP)I homolog (α-COPI), tagged with GFP, was used to investigate the role(s) of fungal Golgi in polar growth. α-COPI-GFP co-localized with the known Golgi marker, α-2,6-sialyltransferase (ST), tagged with red fluorescent protein (RFP), in untreated hyphae. Based on this observation, I propose that α-COPI-GFP can be used as a proxy for fungal Golgi localization. Fungal Golgi were more abundant at hyphal tips than subapically. Fungal Golgi forward (tipward) velocity correlated with hyphal growth rate. Fungal Golgi forward velocity was, on average, approximately ten times greater than average hyphal growth rate. Actin depolymerization reduced fungal Golgi forward velocity while MT depolymerization did not. However, MT stabilization increased fungal Golgi forward velocity. <p>Polymerized MTs do not appear to be essential for hyphal growth but do appear to be involved in hyphal growth rate variability. MTs also appear to play some role in the movement of fungal Golgi. The distribution and movement of fungal Golgi is clearly related to polarity.

Traditionally, hydrogen sulfide (H2S) has been regarded as toxic. It can affect the various human systems and even cause death. However, research in the 1990s has shown that H2S can be endogenously generated by many cells and tissues in mammalian bodies, and that H2S also may play physiological roles such as those of neuromodulator and vasorelaxant in the biological system. As such, the precise measurement of the amount of H2S in a mammalian body has generated researchers strong interest. The ultimate goal of such a measurement should be conducted in-vivo and in real time.<p>The existing methods for H2S measurement require both a large quantity of tissue samples and a complex procedure, so they are not highly practicable for the purpose of achieving the aforementioned goal. In this dissertation, a new method that uses carbon nanotube as an absorbent or transducer and laser-based microscopy techniques (Raman and confocal laser scanning microscopy) as signal excitation and acquisition is proposed and developed. Experimental studies are described of using this new method for analysis of both distilled water samples and serum samples in which a group of proteins are present. <p>The study concludes that the new method (1) can measure H2S in water solutions down to a low level of concentration of 10 µM, (2) can measure H2S in sera down to a low concentration of approximately 20 µM), and (3) has a high feasibility for being used in the clinical context. Regarding (3), this is confirmed by presenting a control system that allows the laser microscopy to track carbon nanotube in a solution that has Brownian motion.<p>While not having reached the ultimate goal as mentioned above, this work advances the state-of-the-art of the measurement of low concentration and nano-quantity of H2S in water and serum samples, in particular providing a promise toward a real-time and in-vivo H2S measurement.

Filamentous fungi, such as <i>Aspergillus nidulans</i>, are composed of tubular, highly polarized, multinucleate cells called hyphae. Polar growth involves secretion specifically at the hyphal tip. Secretion involves intracellular transport and co-ordination of the cytoskeleton and the endomembrane system. <p>Intracellular transport is likely mediated by cytoskeletal elements, which, in fungal cells consist primarily of actin and microtubules (MTs). An <i>A. nidulans</i> strain transformed with green fluorescent protein (GFP) tagged α-tubulin was utilized in the investigation of relationship between cytoplasmic MT arrays and hyphal growth rate. <i>A. nidulans</i> MTs were observed to be long and flexuous and to run roughly parallel to the long axis of hyphae. No correlation between relative MT abundance and hyphal growth rate was observed, although non-growing hyphae had a lower relative MT abundance than growing hyphae. Actin depolymerization decreased hyphal growth rate while MT depolymerization did not. MT stabilization increased hyphal growth rate. Ethanol, the solvent in which the MT and actin inhibitors were dissolved, increased both average overall growth rate and growth rate variability for individual hyphae. Taxol appeared to interact with irradiation to decreased growth rate during imaging. <p>Golgi are involved in secretion and potentially in polar growth. An <i>A. nidulans</i> α-coatomer protein (COP)I homolog (α-COPI), tagged with GFP, was used to investigate the role(s) of fungal Golgi in polar growth. α-COPI-GFP co-localized with the known Golgi marker, α-2,6-sialyltransferase (ST), tagged with red fluorescent protein (RFP), in untreated hyphae. Based on this observation, I propose that α-COPI-GFP can be used as a proxy for fungal Golgi localization. Fungal Golgi were more abundant at hyphal tips than subapically. Fungal Golgi forward (tipward) velocity correlated with hyphal growth rate. Fungal Golgi forward velocity was, on average, approximately ten times greater than average hyphal growth rate. Actin depolymerization reduced fungal Golgi forward velocity while MT depolymerization did not. However, MT stabilization increased fungal Golgi forward velocity. <p>Polymerized MTs do not appear to be essential for hyphal growth but do appear to be involved in hyphal growth rate variability. MTs also appear to play some role in the movement of fungal Golgi. The distribution and movement of fungal Golgi is clearly related to polarity.

Traditionally, hydrogen sulfide (H2S) has been regarded as toxic. It can affect the various human systems and even cause death. However, research in the 1990s has shown that H2S can be endogenously generated by many cells and tissues in mammalian bodies, and that H2S also may play physiological roles such as those of neuromodulator and vasorelaxant in the biological system. As such, the precise measurement of the amount of H2S in a mammalian body has generated researchers strong interest. The ultimate goal of such a measurement should be conducted in-vivo and in real time.<p>The existing methods for H2S measurement require both a large quantity of tissue samples and a complex procedure, so they are not highly practicable for the purpose of achieving the aforementioned goal. In this dissertation, a new method that uses carbon nanotube as an absorbent or transducer and laser-based microscopy techniques (Raman and confocal laser scanning microscopy) as signal excitation and acquisition is proposed and developed. Experimental studies are described of using this new method for analysis of both distilled water samples and serum samples in which a group of proteins are present. <p>The study concludes that the new method (1) can measure H2S in water solutions down to a low level of concentration of 10 µM, (2) can measure H2S in sera down to a low concentration of approximately 20 µM), and (3) has a high feasibility for being used in the clinical context. Regarding (3), this is confirmed by presenting a control system that allows the laser microscopy to track carbon nanotube in a solution that has Brownian motion.<p>While not having reached the ultimate goal as mentioned above, this work advances the state-of-the-art of the measurement of low concentration and nano-quantity of H2S in water and serum samples, in particular providing a promise toward a real-time and in-vivo H2S measurement.

Many Americans are afflicted with inflammation of the colon. They are also at a higher risk of developing colon cancer. Confocal microscopy of bulk epithelial tissue has the potential to provide information on tissue structural properties that may be lost in the fixation and slicing procedures required for histopathology. Optical sectioning provides images in three dimensions capturing the organizational structure of cells and colon crypts throughout the entire colon. I have constructed a custom built fluorescence and reflectance confocal microscope for imaging molecular and morphological changes associated with development of inflammation in a mouse model. A confocal microscope is a point scanning system that removes out of focus light by placing a pinhole aperture in the conjugate image plane located in front of the detector. We have two sources, 488 nm and 811 nm, for fluorescence and reflectance imaging, respectively. A polygon scanning mirror and a galvanometer scanning mirror allow for a variable scan rate between 8 and 15 fps. The lateral resolution of the system is approximately 3 μm with an axial resolution of 6 μm and 4 μm for reflectance and fluorescence mode, respectively. As colon tissue becomes inflamed, there is a distinct change in the structure and architecture of the tissue. The colon crypts are no longer uniform in size or distribution throughout the tissue. Having a large field of view of 1mm2 allows for many colon crypts to be visualized within a single frame. Histology was performed on the same tissue imaged for the inflammatory study confirming the constructed confocal microscope’s ability to characterize inflamed tissue and the potential use for guided biopsy. Mosaicing, or image tiling, is an imaging technique that stitches single frames together to produce a much larger field of view. An extended frame with 1 mm x 2 cm field of view is achieved within seconds. This extended frame would allow mosaicing of the entire mouse colon much faster than conventional methods without loss of resolution. The acquired confocal images of colon tissue demonstrate the microscope’s ability to resolve cell nuclei lining the colon crypts within a relatively large field of view.

Thesis (M. S.. in Biomedical Engineering)--Vanderbilt University, Dec. 2009. / Title from title screen. Includes bibliographical references.

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