Nonlinear Optical Imaging and Microprocessing to Study SH3GLB2 Protein Polymerization

• Main Authors: Yong-DaSie, 謝永達
• Other Authors: Shean-jen Chen
• Format: Others
• Language: en_US
• Published: 2012
• Online Access: http://ndltd.ncl.edu.tw/handle/13994686851826525597

碩士 === 國立成功大學 === 工程科學系碩博士班 === 100 === In this thesis, an ultrafast laser system with three-dimensional (3D) molecular imaging and microprocessing has been utilized to investigate SH3GLB2 protein polymerization. In addition, nano-optics theory and bio-molecular probe technique are implemented. In humans, the Spas-1 ortholog SH3GLB2 has been reported to be overexpressed in prostate cancer metastases. It is observed that in the SH3GLB2 overexpressed African green monkey kidney (COS-7 fibroblasts) cell lines the endophilin B2 protein can aggregate to form clusters around the cell nucleus. The research content of this paper can be divided into three parts. The first part is focused on improving the path planning of a developed Ti:sapphire femtosecond laser system. A Visual Studio C++ program having stereo lithography format file transformation and vector scan path allocation has been developed as a DLL (dynamic link library) format and integrated into the LabVIEW platform of the laser system. Through the above integration, the laser microprocessing can be directly manipulated from the computer aid design of 3D microstructures into the laser processing terminal. In the second part, the developed system was utilized to investigate the recovery of SH3GLB2 protein polymerization clusters after photon damage via fluorescence recovery after photobleaching technique based on observing the protein fluorescence variation with time via single photon counting technique. Optical flow method and sum of absolute difference method were combined as a tracking algorithm in two-photon time lapse imaging which evaluates the effective speed of the protein clusters in cytoplasm. The experimental results are used to study and investigate the TGF-beta1-regulated SH3GLB2 protein assembly and the relation to the ATP metabolism since it is always need energy to participate in when the gene express or protein transport. Finally, nano-optics theory and fluorescence lifetime image microscopy are utilized to study the binding affinity of SH3GLB2 expression construct transfected with a green fluorescent protein (GFP)-tagged and red fluorescence protein (DsRed).