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Quantitative determination of individual urinary glycosaminoglycans in mucopolysaccharidosis by enzymes.January 1998 (has links)
submitted by Chair Siu Fan. / Thesis (M.Sc.)--Chinese University of Hong Kong, 1998. / Includes bibliographical references (leaves 75-83). / Chapter 1 --- INTRODUCTION --- p.1 / Chapter 2 --- LITERATIRE REVIEW --- p.3 / Chapter 2.1 --- Causes and clinical syndromes in MPS --- p.3 / Chapter 2.1.1 --- MPS --- p.4 / Chapter 2.1.2 --- Classification of MPS --- p.4 / Chapter 2.1.2.1 --- MPS I (Hurler's syndrome) --- p.5 / Chapter 2.1.2.2 --- MPS IS (Scheie syndrome) --- p.5 / Chapter 2 .1.2.3 --- MPS II (Hunter's disease) --- p.6 / Chapter 2.1.2.4 --- MPS Type III (The Sanfilippo diseases) --- p.6 / Chapter 2.1.2.5 --- MPS Type IV (Morquio's disease) --- p.6 / Chapter 2.1.2.6 --- MPS Type VI (Maroteaux 一 Lamy syndrome) --- p.7 / Chapter 2.1.2.7 --- MPS Type VII (Sly syndrome) --- p.8 / Chapter 2.1.3 --- Treatment and prospects for MPS --- p.8 / Chapter 2.1.3.1 --- To manage the handicaps and disabilities --- p.8 / Chapter 2.1.3.2 --- Enzyme replacement --- p.9 / Chapter 2.1.3.3 --- Bone marrow transplantation --- p.10 / Chapter 2.2 --- Basic aspects of GAG --- p.10 / Chapter 2.2.1 --- Distributions of GAG --- p.12 / Chapter 2.2.2 --- Functions and Roles of GAG --- p.12 / Chapter 2.2.3 --- Stepwise degradation of GAGs --- p.12 / Chapter 2.2.4 --- Source of urinary GAG --- p.13 / Chapter 2.2.5 --- Common features of GAGS --- p.14 / Chapter 2.2.6 --- Factors affecting the excretion pattern ot GAG --- p.16 / Chapter 2.3 --- Methods for MPS Diagnosis --- p.16 / Chapter 2.3.1 --- Qualitative urine methods for screening and typing --- p.16 / Chapter 2.3.1.1 --- Spot tests --- p.16 / Chapter 2.3.1.2 --- Precipitation methods --- p.16 / Chapter 2.3.1.3 --- One-dimensional electrophoresis --- p.17 / Chapter 2.3.1.4 --- Two-dimensional electrophoresis --- p.17 / Chapter 2.3.1.5 --- Thin layer chromatography --- p.17 / Chapter 2.3.2 --- Quantitative methods for urinary GAG --- p.17 / Chapter 2.3.2.1 --- Measurement of hexuronic acid --- p.18 / Chapter 2.3.2.2 --- HPLC or Column chromatography --- p.18 / Chapter 2.3.2.3 --- Dye-binding methods --- p.19 / Chapter 2.3.3 --- Cytological studies --- p.19 / Chapter 2.3.4 --- Tissue culture --- p.20 / Chapter 2.3.5 --- Tissue biopsy --- p.20 / Chapter 2.3.6 --- Prenatal diagnosis of the MPS --- p.21 / Chapter 2.4 --- Bacterial GAG hydrolytic enzymes --- p.23 / Chapter 2.5 --- Summary of Literature Review --- p.25 / Chapter 3. --- AIMS OF STUDY --- p.26 / Chapter 4. --- MATERIALS AND METHODS --- p.26 / Chapter 4.1 --- Sample collection --- p.26 / Chapter 4.2 --- Materials &-Equipment --- p.26 / Chapter 4.3 --- Preparation of Reagents and Standards --- p.27 / Chapter 4.3.1 --- Stock DMB reagent solutions --- p.27 / Chapter 4.3.2 --- Working DMB solution --- p.27 / Chapter 4.3.3 --- GAG standards --- p.27 / Chapter 4.3.4 --- Reagents for electrophoresis --- p.27 / Chapter 4.3.4.1 --- 0.1M barium acetate solution --- p.27 / Chapter 4.3.4.2 --- 15% ethanolic barium acetate --- p.28 / Chapter 4.3.4.3 --- 50% ethanolic barium acetate --- p.28 / Chapter 4.3.4.4 --- Alcian blue working solution --- p.28 / Chapter 4.3.4.5 --- 0.1M Tris Buffer --- p.28 / Chapter 4.3.4.6 --- CTB Tris solution --- p.28 / Chapter 4.3.4.7 --- 2.0M lithium chloride --- p.28 / Chapter 4.3.5 --- Reagents for enzymatic degradation --- p.28 / Chapter 4.3.5.1 --- Reconstitution of CSE enzyme --- p.28 / Chapter 4.3.5.2 --- Reconstitution of DSE enzyme --- p.29 / Chapter 4.3.5.3 --- Reconstitution ofHSE I enzyme --- p.29 / Chapter 4.4 --- Methods --- p.31 / Chapter 4.4.1 --- Cobas Bio DMB method --- p.31 / Chapter 4.4.2 --- Cobas Fara DMB method --- p.31 / Chapter 4.4.3 --- Evaluation of methods --- p.31 / Chapter 4.4.3.1 --- To study the matrix effect --- p.31 / Chapter 4.4.3.2 --- Calibration --- p.31 / Chapter 4.4.3.3 --- Precision performance --- p.34 / Chapter 4.4.3.4 --- Linearity check --- p.34 / Chapter 4.4.3.5 --- Detection Limit --- p.34 / Chapter 4.4.3.6 --- Recovery study --- p.35 / Chapter 4.4.3.7 --- Correlation with Cobas Bio to develop the reference range --- p.35 / Chapter 4.4.4 --- Electrophoresis method --- p.36 / Chapter 4.4.4.1 --- Sample preparation --- p.36 / Chapter 4.4.4.2 --- Electrophoresis procedure --- p.36 / Chapter 4.4.5 --- Enzymatic degradation method --- p.37 / Chapter 4.4.5.1 --- Digestion of GAG in aqueous and urine matrix --- p.37 / Chapter 4.4.5.2 --- To optimize the amount of enzyme used to degrade GAG --- p.38 / Chapter 4.4.5.3 --- To study the specificity of GAG degrading enzyme --- p.39 / Chapter 4.4.5.4 --- To study the interaction of GAG --- p.40 / Chapter 4.4.5.5 --- To study the stability of enzyme CSE and DSE --- p.40 / Chapter 4.4.5.6 --- Study MPS patient sample --- p.41 / Chapter 5 --- Results --- p.42 / Chapter 5.1 --- Performance characteristics of the DMB method --- p.42 / Chapter 5.1.1 --- Matrix effect --- p.42 / Chapter 5.1.2 --- Calibration --- p.42 / Chapter 5.1.3 --- Precision performance --- p.42 / Chapter 5.1.4 --- Linearity Range --- p.42 / Chapter 5.1.5 --- Detection limit --- p.42 / Chapter 5.1.6 --- Recovery --- p.47 / Chapter 5.1.7 --- Correlation of Cobas Fara with Cobas Bio --- p.47 / Chapter 5.2 --- Results of GAG enzymatic degradation --- p.50 / Chapter 5.2.2 --- To optimise the amount of enzyme for GAG degradation --- p.57 / Chapter 5.2.3 --- The specificity of GAG degrading enzymes --- p.57 / Chapter 5.2.4 --- The interaction of GAG --- p.57 / Chapter 5.2.5 --- The stability of enzymes --- p.57 / Chapter 5.2.6 --- MPS patient study --- p.57 / Chapter 5.2.6.1 --- Type I/II/VI/VII --- p.57 / Chapter 5.2.6.2 --- MPS Type III patient 1 --- p.64 / Chapter 5.2.6.3 --- MPS Type IIIC patient 2 --- p.64 / Chapter 6. --- DISCUSSION --- p.67 / Chapter 6.1 --- Automated DMB method on Cobas Fara --- p.67 / Chapter 6 2 --- GAG specific degradation enzymes --- p.70 / Chapter 7. --- CONCLUSION & SUGGESTION FOR FUTURE STUDIES --- p.73 / Chapter 8. --- REFERENCES --- p.75
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