Chan, Ka Ying. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 133-150). / Abstract also in Chinese. / ABSTRACT --- p.i / 摘要 --- p.iii / ACKNOWLEDGEMENTS --- p.iv / CONTRIBUTORS --- p.vi / TABLE OF CONTENTS --- p.vii / LIST OF TABLES --- p.x / LIST OF FIGURES --- p.xi / LIST OF ABBREVIATIONS --- p.xiii / Chapter SECTION I: --- BACKGROUND --- p.1 / Chapter CHAPTER 1: --- PRENATAL DIAGNOSIS OF FETAL TRISOMY 21 --- p.2 / Chapter 1.1 --- Down syndrome --- p.2 / Chapter 1.2 --- Current methods of prenatal diagnosis of fetal trisomy 21 --- p.3 / Chapter 1.2.1 --- Non-invasive procedures --- p.3 / Chapter 1.2.2 --- Invasive procedures --- p.5 / Chapter 1.3 --- Alternative methods for the prenatal diagnosis of fetal trisomy 21 --- p.7 / Chapter CHAPTER 2: --- CELL-FREE FETAL NUCLEIC ACIDS IN MATERNAL PLASMA --- p.13 / Chapter 2.1 --- Circulating fetal cells --- p.15 / Chapter 2.2 --- Circulating cell-free fetal nucleic acids --- p.15 / Chapter 2.3 --- Diagnostic applications of cell-free fetal nucleic acids in maternal plasma --- p.17 / Chapter 2.4 --- Digital relative chromosome dosage approach --- p.20 / Chapter 2.5 --- Validation of digital RCD approach on artificial DNA mixtures --- p.22 / Chapter SECTION II --- : MATERIALS AND METHODS --- p.25 / Chapter CHAPTER 3: --- QUANTITATIVE ANALYSIS OF NUCLEIC ACIDS --- p.26 / Chapter 3.1 --- Subject recruitment and sample collection --- p.26 / Chapter 3.2 --- Sample processing --- p.26 / Chapter 3.3 --- Nucleic acid extraction --- p.27 / Chapter 3.3.1 --- Extraction of DNA from placental tissues --- p.27 / Chapter 3.3.2 --- Extraction of DNA from maternal blood cells --- p.27 / Chapter 3.4 --- Matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS) --- p.28 / Chapter 3.5 --- Paralogous sequence assays optimisation workflow --- p.31 / Chapter 3.5.1 --- Monoplex paralogous sequence assays --- p.31 / Chapter 3.5.2 --- Multiplex paralogous sequence assay --- p.38 / Chapter 3.6 --- Digital PCR --- p.42 / Chapter 3.6.1 --- Principle --- p.42 / Chapter 3.6.2 --- Digital multiplex paralogous sequence assay --- p.42 / Chapter 3.7 --- Statistical analysis --- p.46 / Chapter 3.7.1 --- Disease classification of samples --- p.46 / Chapter 3.7.2 --- Poisson distribution --- p.46 / Chapter 3.7.3 --- Data analysis --- p.48 / Chapter 3.7.4 --- Sequential probability ratio test (SPRT) analysis --- p.49 / Chapter SECTION III: --- ASSAY DEVELOPMENT --- p.53 / Chapter CHAPTER 4: --- TESTING OF ASSAY SPECIFICITY WITH CORIELL CELL LINES --- p.54 / Chapter 4.1 --- Coriell cell lines --- p.54 / Chapter 4.2 --- Specificity of initial PCR primers --- p.56 / Chapter 4.2.1 --- Principle --- p.56 / Chapter 4.2.2 --- Materials and methods --- p.56 / Chapter 4.2.3 --- Results --- p.60 / Chapter 4.2.4 --- Conclusion --- p.63 / Chapter 4.3 --- Specificity of the iPLEX® Gold extension primers --- p.63 / Chapter 4.3.1 --- Principle --- p.63 / Chapter 4.3.2 --- Materials and methods --- p.64 / Chapter 4.3.3 --- Results --- p.65 / Chapter 4.4 --- Further analysis on the specificity of PV2107a initial PCR primers --- p.67 / Chapter 4.5 --- Conclusion --- p.71 / Chapter CHAPTER 5: --- ASSAY OPTIMISATION --- p.72 / Chapter 5.1 --- Introduction --- p.72 / Chapter 5.2 --- Optimisation of initial PCRs with AmpliTaq Gold® DNA polymerase followed by homogeneous MassEXTEN´DёØ (hME) assays (Sequenom) --- p.72 / Chapter 5.2.1 --- Optimisation of initial PCR reactions --- p.72 / Chapter 5.2.2 --- Principle of homogeneous MassEXTEN´DёØ assays (Sequenom)… --- p.75 / Chapter 5.2.3 --- Homogeneous MassEXTEN´DёØ assays (Sequenom) on euploid and T21 samples --- p.76 / Chapter 5.3 --- Assay selection by iPLEX® Gold single base primer extension reactions (Sequenom) --- p.82 / Chapter 5.4 --- Optimisation of multiplex PCR with AmpliTaq Gold® DNA polymerase --- p.88 / Chapter 5.5 --- Optimisation of multiplex iPLEX® Gold single base primer extension reaction --- p.93 / Chapter 5.6 --- Single molecule detection test for the multiplex paralogous sequence assays … --- p.103 / Chapter SECTION IV: --- ANALYSIS OF CLINICAL SAMPLES --- p.107 / Chapter CHAPTER 6: --- DISEASE CLASSIFICATION OF EUPLOID AND TRISOMY SAMPLES WITH MULTIPLEX PARALOGOUS SEQUENCE ASSAY --- p.108 / Chapter 6.1 --- Introduction --- p.108 / Chapter 6.2 --- Materials and methods --- p.109 / Chapter 6.2.1 --- Sample collection --- p.109 / Chapter 6.2.2 --- Experimental design --- p.110 / Chapter 6.3 --- Results --- p.111 / Chapter 6.4 --- Discussion --- p.114 / Chapter SECTION V: --- CONCLUDING REMARKS --- p.122 / Chapter CHAPTER 7: --- CONCLUSION AND FUTURE PERSPECTIVES --- p.123 / Chapter 7.1 --- Conclusion --- p.123 / Chapter 7.2 --- Future perspectives --- p.124 / Appendix 1 --- p.126 / Appendix II --- p.127 / REFERENCE --- p.133

published_or_final_version / Obstetrics and Gynaecology / Master / Master of Philosophy

Bibliography: leaves 223-229. / xiii, 229 leaves : ill. ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / This thesis considers several aspects of multivariate analysis motivated by the South Australian prenatal Down's syndrome screen, which seeks to identify those pregnancies at high risk to Down's syndrome from several risk factors, amongst them maternal age and measurements from four analyte concentrations found in maternal blood. The primary aim is to discriminate between the unaffected and affected pregnancies in terms of these observed risk factors. Through the development of multivariate regression models it is examined how the Mahalanobis distance might be decomposed into distances that give more insight into where the discrimination is derived and how this might change with the gestational age at which the analyte concentrations are measured. A second multivariate aspect motivated by the screen surrounds the detection of directional outliers and the incorporation of knowledge of likely departures into the outlier identification technique. A method is also developed to account for misreporting in inferences of gestational age. Similarities with measurement error and misclassification models are explored. / Thesis (Ph.D.)--University of Adelaide, Dept. of Statistics, 1998?