MOLECULAR BIOLOGY AND BIOTECHNOLOGY

By JOSHUA SMITH

CONTENTS PART I CELL LINE DEVELOPMENT AND SELECTION 1 HIGH THROUGHPUT SYNCHRONIZATION OF MAMMALIAN CELL CULTURES BY SPIRAL MICROFLUIDICS 3 2 INSECT CELL LINE DEVELOPMENT USING FLP MEDIATED CASSETTE EXCHANGE TECHNOLOGY 15 3 SCALABLE TRANSIENT GENE EXPRESSION IN ADHERENT MAMMALIAN CELLS USING POLYETHYLENIMINE 29 4 SCALABLE TRANSIENT PROTEIN EXPRESSION 35 5 DNA PROFILING AND CHARACTERIZATION OF ANIMAL CELL LINES 57 PART II TECHNIQUES FOR PROCESS DEVELOPMENT 6 TOOLS FOR HIGH THROUGHPUT PROCESS AND MEDIUM OPTIMIZATION 77 7 DESIGNING MEDIA FOR ANIMAL CELL CULTURE CHO CELLS THE INDUSTRIAL STANDARD 89 8 FEED OPTIMIZATION IN FED BATCH CULTURE 105 9 SCREENING AND OPTIMIZATION OF CHEMICALLY DEFINED MEDIA AND FEEDS WITH INTEGRATED AND STATISTICAL APPROACHES 117 10 EVALUATION OF SOLID AND POROUS MICROCARRIERS FOR CELL GROWTH AND PRODUCTION OF RECOMBINANT PROTEINS 137 11 MICROBIOREACTORS AND SCALE DOWN MODELS GROWTH OF CHO CELLS USING THE PALL MICRO24 MICROREACTOR SYSTEM 149 PART III CELL CHARACTERISATION AND ANALYSIS 12 MONITORING CELL GROWTH VIABILITY AND APOPTOSIS 169 13 METABOLIC FLUX ESTIMATION IN MAMMALIAN CELL CULTURES 193 14 QUENCHING METHODS FOR THE ANALYSIS OF INTRACELLULAR METABOLITES 211 15 NMR METHODS FOR METABOLOMICS OF MAMMALIAN CELL CULTURE BIOREACTORS 223 16 1H NMR PROTOCOL FOR EXOMETABOLOME ANALYSIS OF CULTURED MAMMALIAN CELLS 237 PART IV CULTIVATION TECHNIQUES 17 MICROFLUIDIC PERFUSION CULTURE 251 18 DISPOSABLE BIOREACTORS FOR INOCULUM PRODUCTION AND PROTEIN EXPRESSION 265 19 CONFIGURATION OF BIOREACTORS 285 20 PROCESS ANALYTICAL TECHNOLOGY PAT IN INSECT AND MAMMALIAN CELL CULTURE PROCESSES DIELECTRIC SPECTROSCOPY AND FOCUSED BEAM REFLECTANCE MEASUREMENT FBRM 313 21 CELL ASSESSMENT BY AT LINE MICROSCOPY 343 22 SEED TRAIN OPTIMIZATION FOR CELL CULTURE 355 PART V SPECIAL APPLICATIONS 23 VACCINE PRODUCTION UPSTREAM PROCESSING 24 EFFICIENT PROCESSES FOR PROTEIN EXPRESSION USING RECOMBINANT BACULOVIRUS PARTICLES 395 25 CHROMATOGRAPHIC TECHNIQUES IN THE DOWNSTREAM PROCESSING OF PROTEINS IN BIOTECHNOLOGY 419 26 VIRUS SEPARATION USING MEMBRANES 459 1 PART I CELL LINE DEVELOPMENT AND SELECTION 1