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Yim Ka Wing. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 93-102). / Abstracts in English and Chinese. / Statement --- p.I / Abstract --- p.Ii / Acknowledgements --- p.Iv / Publications and awards --- p.Vii / General abbreviations --- p.Viii / Technical abbreviations --- p.Ix / Abbreviations of chemicals --- p.X / List of figures --- p.Xi / List of tables --- p.Xiii / Table of contents --- p.Xiv / Chapter Chapter 1 --- Introduction --- p.1 / Chapter Chapter 2 --- Pre-eclampsia --- p.3 / Chapter 2.1 --- Inadequate Uteroplacental Circulation --- p.5 / Chapter 2.2 --- Placenta Ischaemia --- p.7 / Chapter 2.3 --- Oxidative Stress --- p.7 / Chapter 2.4 --- Systemic Inflammatory Response --- p.9 / Chapter 2.5 --- The Continuum Theory of Clinical Syndromes --- p.12 / Chapter 2.6 --- Origin of Stimuli to Inflammatory Response --- p.13 / Chapter Chapter 3 --- Proteomic Analysis --- p.16 / Chapter 3.1 --- Methodology in Proteomic Research --- p.17 / Chapter 3.1.1 --- 2D-PAGE --- p.17 / Chapter 3.1.2 --- Mass Spectrometry --- p.18 / Chapter 3.1.2.1 --- Peptide Mass Fingerprinting --- p.18 / Chapter 3.1.2.2 --- Product-ion Data --- p.19 / Chapter 3.1.2.3 --- Matrix-assisted Laser Desorption/Ionization Tandem Time-of-Flight Mass Spectrometry (MALDI-TOF/TOF MS) --- p.19 / Chapter 3.1.3 --- Bioinformatics Tools --- p.20 / Chapter 3.2 --- Applications of Proteomic Analysis --- p.20 / Chapter Chapter 4 --- Materials and Methods --- p.21 / Chapter 4.1 --- Overview --- p.21 / Chapter 4.2 --- Patients --- p.23 / Chapter 4.2.1 --- Pre-eclampsia --- p.23 / Chapter 4.2.2 --- Recruitment --- p.23 / Chapter 4.3 --- Stage I: Comparative Proteomic Analysis --- p.24 / Chapter 4.3.1 --- Sample Processing --- p.24 / Chapter 4.3.2 --- Modified Bradford's Method --- p.24 / Chapter 4.3.3 --- Albumin Depletion --- p.25 / Chapter 4.3.4. --- Comparative Proteomic Analysis --- p.26 / Chapter 4.3.4.1 --- First Dimension: Isoelectric Focusing (IEF) --- p.28 / Chapter 4.3.4.2 --- Reduction and Alkylation --- p.29 / Chapter 4.3.4.3 --- Second Dimension: SDS-PAGE --- p.30 / Chapter 4.3.4.4 --- Gel Imaging --- p.31 / Chapter 4.3.5 --- 2D-PAGE Data Analysis --- p.32 / Chapter 4.3.5.1 --- Gaussian Spot --- p.32 / Chapter 4.3.5.2 --- Matchset --- p.33 / Chapter 4.3.5.3 --- Normalization --- p.33 / Chapter 4.3.5.4 --- Significance Analysis of Microarrays --- p.34 / Chapter 4.4 --- Stage II: Protein Identification --- p.35 / Chapter 4.4.1 --- Tryptic Peptide Fingerprinting --- p.37 / Chapter 4.4.1.1 --- Removal of Silver Ions --- p.37 / Chapter 4.4.1.2 --- Reduction and Alkylation --- p.38 / Chapter 4.4.1.3 --- In-gel Digestion --- p.38 / Chapter 4.4.1.4 --- Clean up of Peptides --- p.39 / Chapter 4.4.1.5 --- Mass Spectrometric Analysis --- p.39 / Chapter 4.4.2 --- Database search: Profound-Peptide Mapping --- p.40 / Chapter 4.4.3 --- Database search: Mascot-MS/MS Ion Search --- p.40 / Chapter 4.5 --- Stage III: Immunoassays --- p.41 / Chapter 4.6 --- Sample Size and Power Calculations --- p.42 / Chapter 4.7 --- Statistical Analysis --- p.42 / Chapter Chapter 5 --- Results --- p.43 / Chapter 5.1 --- Patients' Demographic Characteristics and Obstetric Outcomes for 2D-PAGE Analysis --- p.43 / Chapter 5.2 --- Plasma Protein Quantity --- p.45 / Chapter 5.3 --- 2D-PAGE --- p.47 / Chapter 5.4 --- SAM Analysis --- p.47 / Chapter 5.5 --- Spot # 5204 and Spot # 6210 --- p.50 / Chapter 5.6 --- Protein Identification --- p.54 / Chapter 5.6.1 --- Identification of Protein Spot # 5204 --- p.54 / Chapter 5.6.2 --- Identification of Protein Spot # 6210 --- p.61 / Chapter 5.7 --- Patients' Demographic Characteristics and Obstetric Outcomes for Immunoassays --- p.68 / Chapter 5.8 --- Immunoassays --- p.70 / Chapter Chapter 6 --- Discussion --- p.81 / Chapter 6.1 --- Role of ficolins and MBL in complement activation pathway --- p.82 / Chapter 6.2 --- Structure of ficolins and MBL --- p.83 / Chapter 6.2.1 --- H-ficolin --- p.85 / Chapter 6.2.2 --- L-ficolin --- p.86 / Chapter 6.3 --- Immune Response and Pre-eclampsia --- p.86 / Chapter 7 --- Conclusion --- p.89 / Appendix --- p.90 / References --- p.93

Liu , Huan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 43-45). / Abstracts in English and Chinese. / Abstract --- p.i / 摘要 --- p.iii / Acknowledgement --- p.iv / Table of contents --- p.vi / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1.1 --- Gel electrophoresis of proteins --- p.1 / Chapter 1.1.1 --- Principles of gel electrophoresis --- p.1 / Chapter 1.1.2 --- Polyacrylamide gel --- p.3 / Chapter 1.1.3 --- Buffer systems --- p.5 / Chapter 1.1.4 --- Capillary electrophoresis --- p.6 / Chapter 1.2 --- Methods to transfer proteins from gel to membrane --- p.7 / Chapter 1.2.1 --- Simple diffusion --- p.8 / Chapter 1.2.2 --- Ultrasound transfer --- p.8 / Chapter 1.2.3 --- Tank transfer --- p.9 / Chapter 1.2.4 --- Semidry transfer --- p.9 / Chapter 1.2.5 --- Transfer efficiency improvements --- p.10 / Chapter 1.3 --- Visualizing immunoblots --- p.11 / Chapter 1.3.1 --- Membrane staining --- p.11 / Chapter 1.3.2 --- Radiometric detection in blotting --- p.12 / Chapter 1.3.3 --- Bioluminescence-enhanced detection --- p.13 / Chapter 1.3.4 --- Chemiluminescence-enhanced detection --- p.14 / Chapter 1.4 --- Miniaturized electrophoresis and blotting methods --- p.15 / Chapter 1.5 --- Objective of the project --- p.18 / Chapter Chapter 2 --- Dip-pen gel electrophoresis in capillaries --- p.20 / Chapter 2.1 --- Introduction --- p.20 / Chapter 2.2 --- Experimental section --- p.21 / Chapter 2.2.1 --- Materials and reagents --- p.21 / Chapter 2.2.2 --- PA gel fabrication in capillary --- p.22 / Chapter 2.2.3 --- Setup for electrophoresis in capillary --- p.23 / Chapter 2.3 --- Results and discussion --- p.24 / Chapter 2.3.1 --- PA gel polymerization quality at tip --- p.24 / Chapter 2.3.2 --- Separation efficiency in capillary --- p.25 / Chapter 2.4 --- Conclusions --- p.27 / Chapter Chapter 3 --- Western blotting by dip-pen electrophoresis --- p.28 / Chapter 3.1 --- Introduction --- p.28 / Chapter 3.2 --- Experimental section --- p.30 / Chapter 3.2.1 --- Materials and reagents --- p.30 / Chapter 3.2.2 --- Protein sample preparation --- p.31 / Chapter 3.2.3 --- Dip-pen electrophoresis based Western blot --- p.31 / Chapter 3.2.4 --- Detection on membrane --- p.32 / Chapter 3.3 --- Results and discussion --- p.33 / Chapter 3.3.1 --- Separation performance on nitrocellulose membrane --- p.33 / Chapter 3.3.2 --- Comparison among different %T PA gel --- p.34 / Chapter 3.3.3 --- SDS-protein complexes capture and immunoblotting --- p.38 / Chapter 3.4 --- Conclusions --- p.39 / Chapter Chapter 4 --- Conclusions --- p.40 / Chapter 4.1 --- Summary --- p.40 / Chapter 4.2 --- Future perspective --- p.41 / References --- p.43

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This thesis describes an immunochemical and immunocytochemical study of the bovine brain S-lOOb protein. The two major forms of the S-100 isoproteins (S-lOOa and S-lOOb) were purified to apparent homogeneity from bovine brain. A polyclonal rabbit antiserum to the S-lOOb protein was prepared. The antiserum was characterized by solid phase immunochemical methods. The S-lOOb derived antiserum displayed a high degree of specificity for S-lOOb, but also crossreacted with the purified S-lOOa protein. The characteristics of the immunochemical reactivity of the antiserum towards these two isoproteins suggests the antiserum has specificity for the 6-subunit of the S-100 proteins. An immunohistochemical analysis of the cellular localization of S-lOOb immunoreactivity was undertaken. In the adult rat brain only the astrocytes were S-lOOb immunoreactive. This conclusion is supported by the morphological characteristics of the immunolabelled cells, as well as the observed co-localization of the immunoreactivities for S-lOOb and the Glial Fibrillary Acidic protein (GFAP), the major protein of the astrocyte intermediate filaments. These two antigens were always found to coexist. The immunolabelling of rat brain astrocytes by the S-lOOb derived antiserum stained the entire cell, yielding more complete morphological detail than is possible with the GFAP immunohistochemistry, which only labels the filamentous glial processes. It is concluded that S-lOOb immunohistochemistry could be of general utility in the investigation of astrocyte morphology. The present results also determine that the as yet unknown biological function(s) of the S-100 proteins must be related to a property of astrocytes. / Medicine, Faculty of / Graduate

Call number: LD2668 .T4 1979 F57 / Master of Science

Call number: LD2668 .T4 1955 M85 / Master of Science

published_or_final_version / Botany / Doctoral / Doctor of Philosophy

published_or_final_version / Computer Science / Master / Master of Philosophy

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