by Paul, Liu Po Lung. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1994. / Includes bibliographical references (leaves 86-94). / ACKNOWLEDGMENT --- p.i / TABLE OF CONTENT --- p.ii / "LIST OF TABLE, FIGURE & PHOTO" --- p.viii / ABSTRACT --- p.x / Chapter CHAPTER ONE : --- INTRODUCTION --- p.1 / Chapter 1.1 --- ALKALINE PHOSPHATASE / Chapter 1.1.1 --- Alkaline Phosphatase Isoenzyme --- p.2 / Chapter 1.1.2 --- The Properties of Alkaline Phosphatases --- p.4 / Chapter 1.1.3 --- Serum Alkaline Phosphatases --- p.6 / Chapter 1.1.3.1 --- Placental Alkaline Phosphatase --- p.7 / Chapter 1.1.3.2 --- Intestinal Alkaline Phosphatase --- p.7 / Chapter 1.1.3.4 --- Skeletal Alkalne Phosphatase --- p.8 / Chapter 1.1.3.5 --- Hepatic Alkaline Phosphatase --- p.8 / Chapter 1.1.3.3 --- Renal Alkaline Phosphatase --- p.9 / Chapter 1.1.3.6 --- Miscellaneous Alkaline Phosphatase --- p.9 / Chapter 1.1.4 --- Problems in Discriminating the Skeletal and Hepatic Alkaline Phosphatase in Serum --- p.11 / Chapter 1.1.5 --- Quantitative measure of the Bone-Specific Alkaline Phosphatase --- p.12 / Chapter 1.1.6 --- Qualitative Detection of ALP isoenzymes --- p.14 / Chapter 1.2 --- OSTEOSARCOMA --- p.17 / Chapter 1.2.1 --- Definition --- p.17 / Chapter 1.2.2 --- Epidemiology and Statistics --- p.17 / Chapter 1.2.3 --- Clinical Presentation --- p.18 / Chapter 1.2.4 --- Radiographic finding --- p.19 / Chapter 1.2.5 --- Staging of Musculoskeletal Neoplasms --- p.20 / Chapter 1.2.6 --- Treatment of osteosarcoma --- p.21 / Chapter 1.2.6.1. --- Chemotherapy in Prince of Wales Hospital --- p.21 / Chapter 1.3 --- PLASMA AND TISSUE ALKALINE PHOSPHATASE IN NORMAL AND NEOPLASTIC CONDITION --- p.23 / Chapter 1.3.1 --- Normal values of plasma alkaline phosphatase --- p.23 / Chapter 1.3.2 --- Clinical use of elevated plasma & tissue alkaline phosphatase level in neoplastic conditions --- p.25 / Chapter 1.3.2.1 --- Helping the Diagnosis of the Osteosarcoma --- p.25 / Chapter 1.3.2.2 --- Monitoring the effect of chemotherapy --- p.26 / Chapter 1.3.2.3 --- Predicting the clinical course --- p.26 / Chapter 1.3.3 --- Qualitative measurement of ALP in plasma and tissue extract of osteosarcoma patient --- p.29 / Chapter 1.4 --- AIM AND SCOPE OF THE PRESENT DISSERATION --- p.30 / Chapter CHAPTER TWO : --- MATERIALS AND METHODS --- p.32 / Chapter 2.1 --- DIFERENT GROUPS OF PATIENTS --- p.33 / Chapter 2.1.1 --- Monitering the plasma bone specific ALP --- p.33 / Chapter 2.1.1.1 --- Osteosarcoma group --- p.33 / Chapter 2.1.1.2 --- Benign bone tumour group --- p.34 / Chapter 2.1.1.3 --- Metastasis group --- p.34 / Chapter 2.1.2 --- Collection of plasma samples preserve of tumor tissue --- p.34 / Chapter 2.2 --- QUANTITATIVE ANALYSIS OF THE PLASMA AND TISSUE BONE SPECIFIC ALKALINE PHOSPHATASE --- p.36 / Chapter 2.2.1 --- Extraction of tissue ALP --- p.36 / Chapter 2.2.1.1. --- Reagent --- p.36 / Chapter 2.2.1.2. --- Homogenization of the bone tissue --- p.36 / Chapter 2.2.1.3. --- Extraction of ALP --- p.37 / Chapter 2.2.2 --- Assay for Bone-specific ALP --- p.38 / Chapter 2.2.2.1. --- Reagents --- p.38 / Chapter 2.2.2.2. --- Procedures --- p.38 / Chapter 2.3 --- QUALITATIVE MEASUREMENT OF ALP ISOENZYME --- p.40 / Chapter 2.3.1 --- Equipment required --- p.40 / Chapter 2.2.2 --- Practical procedure --- p.40 / Chapter 2.3.3.1 --- Gel casting --- p.40 / Chapter 2.3.3.2 --- Sample preparation and application --- p.42 / Chapter 2.3.3.3 --- Electrofocusing --- p.42 / Chapter 2.3.3.4 --- Western blotting of the protein --- p.43 / Chapter 2.3.3.5 --- Detection methods --- p.45 / Chapter 2.4 --- METHOD OF STATISTICAL ANALYSIS --- p.48 / Chapter CHAPTER THREE : --- RESULTS --- p.49 / Chapter 3.1 --- QUANTITATIVE MEASUREMENT OF PLASMA AND TISSUE BONE SPECIFIC ALKALINE PHOSPHATASE --- p.50 / Chapter 3.1.1 --- General Information of the patients monitoring --- p.50 / Chapter 3.1.2 --- Pretreatment evaluation --- p.52 / Chapter 3.1.3 --- Correlation between the pretreatment plasma ALP levels and prognosis in the osteosarcoma patient group --- p.57 / Chapter 3.1.4 --- "Correlation between the pre-operational, post- operational plasma ALP levels and the prognosis of osteosarcoma" --- p.59 / Chapter 3.1.5 --- Analysis of plasma ALP levels at the time of relapse in osteosarcoma patients --- p.61 / Chapter 3.1.6 --- Usefulness of the plasma ALP levels for monitoring the effectiveness of chemotherapy --- p.62 / Chapter 3.1.7 --- Correlation between the ALP levels in the tumor extract and the prognosis of the osteosarcoma --- p.64 / Chapter 3.2 --- QUALITATIVE ANALYSIS OF THE PLASMA AND TISSUE ALKALINE PHOSPHATASE LEVEL --- p.67 / Chapter 3.2.1 --- Comparison of the result of Isoelectric focusing of the plasma ALP of the osteosarcoma patients and the normal subjects --- p.67 / Chapter 3.2.1 --- Result of Isoelectric focusing of the ALP isoenzymes in the tissue extract of the osteosarcoma and normal bone --- p.70 / Chapter CHAPTER FOUR : --- DISCUSSION --- p.72 / Chapter 4.1 --- USE OF QUANTITATIVE MONITORING OF PLASMA ALP AND MEASURING TISSUE ALP IN OSTEOSARCOMA PATIENTS --- p.73 / Chapter 4.2 --- ISOELECTRIC FORCUSING AS A TECNIQUE FOR QUALITATIVE MEASUREMENT OF PLASMA AND TISSUE ALKALINE PHOSPHATASE --- p.80 / Chapter CHAPTER FIVE : --- CONCLUSION --- p.83 / Chapter CHAPTER SIX : --- BIBILOGRAPHY --- p.85

by Chak Chi Wai. / Thesis (M.Phil)--Chinese University of Hong Kong, 1992. / Includes bibliographical references (leaves 142-155). / ACKNOWLEDGMENTS --- p.i / TABLE OF CONTENT --- p.ii / LIST OF ABBREVIATION --- p.viii / ABSTRACT --- p.x / Chapter CHAPTER ONE: --- INTRODUCTION / Chapter 1.1 --- INTRODUCTION TO ALKALINE PHOSPHATASE --- p.2 / Chapter 1.1.1 --- The Alkaline Phosphatase Isoenzymes --- p.2 / Chapter 1.1.2 --- The Properties of Alkaline Phosphatases --- p.5 / Chapter 1.1.3 --- Serum Alkaline Phosphatases --- p.7 / Chapter 1.1.3.1 --- Intestinal Alkaline Phosphatase --- p.8 / Chapter 1.1.3.2 --- Placental Alkaline Phosphatase --- p.8 / Chapter 1.1.3.3 --- Renal Alkaline Phosphatase --- p.9 / Chapter 1.1.3.4 --- Skeletal Alkaline Phosphatase --- p.9 / Chapter 1.1.3.5 --- Hepatic Alkaline Phosphatase --- p.10 / Chapter 1.1.3.6 --- Miscellaneous Alkaline Phosphatases --- p.10 / Chapter 1.1.4 --- Problems in Discriminating the Skeletal and Hepatic Alkaline Phosphatase in Serum --- p.12 / Chapter 1.1.5 --- Wheat Germ Lectin Precipitation of the Bone- Specific Alkaline Phosphatase --- p.13 / Chapter 1.2 --- STRUCTURE OF BONE AND MECHANISMS OF CALCIFICATION --- p.15 / Chapter 1.2.1 --- Gross Structure of Bone --- p.15 / Chapter 1.2.2 --- The Elements of Bone --- p.17 / Chapter 1.2.2.1 --- Bone Cells --- p.17 / Chapter 1.2.2.2 --- Organic Substances of Bone --- p.19 / Chapter 1.2.2.3 --- Inorganic Substances of Bone --- p.21 / Chapter 1.2.3 --- Mechanisms of Calcification --- p.22 / Chapter 1.3 --- BONE FRACTURE HEALING --- p.24 / Chapter 1.3.1 --- Types of Fracture --- p.24 / Chapter 1.3.2 --- The Process of Bone Fracture Healing --- p.26 / Chapter 1.3.2.1 --- Stage of Hematoma --- p.26 / Chapter 1.3.2.2 --- Stage of Subperiosteal and Endosteal Cellular Proliferation --- p.28 / Chapter 1.3.2.3 --- Stage of Fibrocartilaginous Callus --- p.28 / Chapter 1.3.2.4 --- Stage of Bony Callus --- p.30 / Chapter 1.3.2.5 --- Stage of Remodeling --- p.31 / Chapter 1.4 --- THE OSTEOBLASTIC CHARACTERS OF UMR-106 OSTEOSARCOMA CELL LINE --- p.32 / Chapter 1.4.1 --- Classification of Osteosarcoma --- p.32 / Chapter 1.4.2 --- Derivation of UMR-106 Osteosarcoma Cell Line --- p.33 / Chapter 1.4.3 --- Osteoblastic Characters of UMR-106 --- p.34 / Chapter 1.4.3.1 --- ALP Expression --- p.34 / Chapter 1.4.3.2 --- Hormone Responsive Adenylate Cyclase System --- p.35 / Chapter 1.4.3.3 --- "Cytosolic Receptors for 1,25-Dihydroxy- cholecalciferol" --- p.35 / Chapter 1.5 --- IN VITRO CULTURE OF FETAL RAT CALVARIAL OSTEOBLASTS --- p.37 / Chapter 1.6 --- AIM AND SCOPE OF THIS DISSERTATION --- p.39 / Chapter CHAPTER TWO: --- MATERIALS AND METHODS / Chapter 2.1 --- BONE FRACTURE OPERATION --- p.42 / Chapter 2.1.1 --- Animals --- p.42 / Chapter 2.1.2 --- Blood Sampling and Preparation of Plasma Samples --- p.42 / Chapter 2.1.3 --- Bone Fracture Operation --- p.43 / Chapter 2.1.3.1 --- Reagents and Apparatus --- p.43 / Chapter 2.1.3.2 --- Procedures --- p.44 / Chapter 2.1.4 --- Radiography --- p.50 / Chapter 2.1.5 --- Removal of Tibiae --- p.51 / Chapter 2.1.6 --- Extraction of Callus ALP --- p.51 / Chapter 2.1.6.1 --- Reagent --- p.51 / Chapter 2.1.6.2 --- Homogenization of the Callus --- p.51 / Chapter 2.1.6.3 --- Extraction of ALP --- p.52 / Chapter 2.1.7 --- Assay for Bone-Specific ALP --- p.53 / Chapter 2.1.7.1 --- Reagents --- p.53 / Chapter 2.1.7.2 --- Procedures --- p.54 / Chapter 2.1.8 --- Normal Curve for Plasma Bone-Specific ALP in Rabbits --- p.56 / Chapter 2.1.9 --- The Effects of Tibial Fracture on the Plasma Level of Bone-Specific ALP in Rabbits --- p.56 / Chapter 2.1.10 --- Profile of Plasma Bone-Specific ALP upon a Fracture Healing --- p.57 / Chapter 2.1.11 --- Profile of Callus Bone-Specific ALP at Different Stages of Fracture Healing --- p.57 / Chapter 2.2 --- CLINICAL STUDIES OF PLASMA BONE-SPECIFIC ALP --- p.58 / Chapter 2.2.1 --- Patient Groups --- p.58 / Chapter 2.2.1.1 --- Normal Adults --- p.58 / Chapter 2.2.1.2 --- Fracture Group --- p.58 / Chapter 2.2.1.3 --- Tumor Group --- p.59 / Chapter 2.2.2 --- Assays for Plasma Bone-Specific ALP --- p.59 / Chapter 2.3 --- "IN VITRO CULTURES OF FETAL, RAT OSTEOBLASTS AND UMR-106 OSTEOSARCOMA cell line" --- p.60 / Chapter 2.3.1 --- Animals --- p.60 / Chapter 2.3.2 --- UMR-106 Cell Line --- p.60 / Chapter 2.3.3 --- General Reagents Used for Cell Culture --- p.60 / Chapter 2.3.4 --- Isolation of Calvarial Osteoblasts --- p.64 / Chapter 2.3.4.1 --- Tools and Reagents --- p.64 / Chapter 2.3.4.2 --- Procedures --- p.65 / Chapter 2.3.5 --- Storage of UMR-106 Cell Line --- p.67 / Chapter 2.3.6 --- Subculture of Confluent Monolayer --- p.68 / Chapter 2.3.6.1 --- Reagents --- p.68 / Chapter 2.3.6.2 --- Procedures --- p.69 / Chapter 2.3.7 --- Staining for Calcium Deposits --- p.69 / Chapter 2.3.7.1 --- Reagents --- p.70 / Chapter 2.3.7.2 --- Procedures --- p.70 / Chapter 2.3.8 --- Protein Determination --- p.71 / Chapter 2.3.8.1 --- Reagents --- p.71 / Chapter 2.3.8.2 --- Procedures --- p.71 / Chapter 2.3.9 --- Microdetermination of Inorganic Phosphate --- p.72 / Chapter 2.3.9.1 --- Reagents --- p.72 / Chapter 2.3.9.2 --- Procedures --- p.73 / Chapter 2.3.10 --- Determination of Calcium --- p.73 / Chapter 2.3.10.1 --- Reagent --- p.73 / Chapter 2.3.10.2 --- Procedures --- p.73 / Chapter 2.3.11 --- Extraction and Assay for Cellular ALP --- p.74 / Chapter 2.3.11.1 --- Reagents --- p.74 / Chapter 2.3.11.2 --- Procedures --- p.75 / Chapter 2.3.12 --- Cell Surface ALP Assay --- p.75 / Chapter 2.3.12.1 --- Reagents --- p.75 / Chapter 2.3.12.2 --- Procedures --- p.76 / Chapter 2.3.13 --- Extraction of Calcium Phosphate Deposits --- p.76 / Chapter 2.3.13.1 --- Reagent --- p.76 / Chapter 2.3.13.2 --- Procedures --- p.76 / Chapter 2.3.14 --- Collagen Synthesis Assay --- p.77 / Chapter 2.3.14.1 --- Reagents --- p.77 / Chapter 2.3.14.2 --- Procedures --- p.78 / Chapter CHAPTER THREE: --- EFFECTS OF TIBIAL FRACTURE ON THE LEVEL OF BONE-SPECIFIC ALKALINE PHOSPHATASE IN RABBITS / INTRODUCTION --- p.81 / results: / Chapter 3.1 --- normal curve for plasma bone-specific alp in rabbits --- p.82 / Chapter 3.2 --- THE EFFECTS OF TIBIAL FRACTURE ON THE PLASMA LEVEL OF BONE-SPECIFIC ALP IN RABBITS --- p.84 / Chapter 3.3 --- PROFILE OF THE PLASMA ALP LEVEL UPON HEALING OF TIBIAL FRACTURE --- p.86 / Chapter 3.4 --- RADIOGRAPHY --- p.89 / Chapter 3.5 --- PROFILE OF CALLUS BONE-SPECIFIC ALP ACTIVITY UPON HEALING OF TIBIAL FRACTURE --- p.93 / DISCUSSION --- p.95 / Chapter CHAPTER FOUR: --- CLINICAL STUDIES OF PLASMA BONE-SPECIFIC ALKALINE PHOSPHATASE / INTRODUCTION --- p.100 / RESULTS: / Chapter 4.1 --- NORMAL VALUES --- p.100 / Chapter 4.2 --- FRACTURE GROUP --- p.101 / Chapter 4.3 --- BONE TUMOR GROUP --- p.102 / DISCUSSION --- p.102 / Chapter CHAPTER FIVE: --- IN VITRO CULTURE OF FETAL RAT OSTEOBLASTS AND UMR-106 CELL LINE / INTRODUCTION --- p.105 / RESULTS: / Chapter 5.1 --- IN VITRO MINERALIZATION OF UMR-106 CELLS AND PRIMARY RC CELLS --- p.107 / Chapter 5.2 --- STUDY OF BONE-SPECIFIC ALP RELEASED INTO MEDIUM BY UMR-106 CELLS AND PRIMARY RC CELLS --- p.113 / Chapter 5.3 --- STUDY OF CELLULAR ALP ACTIVITIES AND CALCIUM PHOSPHATE DEPOSITS --- p.116 / Chapter 5.4 --- STUDIES OF CELLULAR ALP ACTIVITIES AND RELATIVE RATES OF COLLAGEN SYNTHESIS --- p.125 / DISCUSSION --- p.128 / Chapter CHAPTER SIX: --- GENERAL DISCUSSION --- p.136 / BIBLIOGRAPHY --- p.142 / APPENDIX --- p.156