Read more
Written for industrial and academic researchers and development scientists in the life sciences industry, Bioprocessing Technology for Production of Biopharmaceuticals and Bioproducts is a guide to the tools, approaches, and useful developments in bioprocessing. This important guide:
* Summarizes state-of-the-art bioprocessing methods and reviews applications in life science industries
* Includes illustrative case studies that review six milestone bio-products
* Discuses a wide selection of host strain types and disruptive bioprocess technologies
List of contents
List of Contributors xi
Part I Case Study 1
1 Bacillus and the Story of Protein Secretion and Production 3
Giulia Barbieri, Anthony Calabria, Gopal Chotani, and Eugenio Ferrari
1.1 Bacillus as a Production Host: Introduction and Historical Account 3
1.2 The Building of a Production Strain: Genetic Tools for B. subtilis Manipulation 5
1.2.1 Promoters 5
1.2.2 Vectors for Building a Production Strain 6
1.2.3 B. subtilis Competent Cell Transformation 7
1.2.4 Protoplasts-Mediated Manipulations 9
1.2.5 Genetics by Electroporation 9
1.3 B. subtilis Secretion Systemand Heterologous Protein Production 9
1.3.1 Bacillus Fermentation and Recovery of Industrial Enzyme 11
1.3.2 Fermentation Stoichiometry 12
1.3.3 Fermentor Kinetics and Outputs 14
1.3.4 Downstream Processing 17
1.4 Summary 21
References 21
2 New Expression Systems for GPCRs 29
Dimitra Gialama, Fragiskos N. Kolisis, and Georgios Skretas
2.1 Introduction 29
2.2 Recombinant GPCR Production - Traditional Approaches for Achieving High-Level Production 39
2.3 Engineered Expression Systems for GPCR Production 42
2.3.1 Bacteria 42
2.3.2 Yeasts 48
2.3.3 Insect Cells 51
2.3.4 Mammalian Cells 54
2.3.5 Transgenic Animals 54
2.3.6 Cell-Free Systems 56
2.4 Conclusion 57
References 58
3 Glycosylation 71
Maureen Spearman, Erika Lattová, Hélène Perreault, andMichael Butler
3.1 Introduction 71
3.2 Types of Glycosylation 72
3.2.1 N-linked Glycans 72
3.2.2 O-linked Glycans 74
3.3 Factors Affecting Glycosylation 76
3.3.1 Nutrient Depletion 76
3.3.2 Fed-batch Cultures and Supplements 79
3.3.3 Specific Culture Supplements 80
3.3.4 Ammonia 82
3.3.5 pH 82
3.3.6 Oxygen 83
3.3.7 Host Cell Systems 83
3.3.8 Other Factors 85
3.4 Modification of Glycosylation 86
3.4.1 siRNA and Gene Knockout/Knockin 86
3.4.2 Glycoprotein Processing Inhibitors and In Vitro Modification of Glycans 88
3.5 Glycosylation Analysis 89
3.5.1 Release of Glycans from Glycoproteins 90
3.5.2 Derivatization of Glycans 91
3.6 Methods of Analysis 91
3.6.1 Lectin Arrays 91
3.6.2 Liquid Chromatography 93
3.6.2.1 HILIC Analysis 93
3.6.2.2 Reversed Phase (RP) and Porous Graphitic Carbon (PGC) Chromatography 95
3.6.2.3 Weak Anion Exchange (WAX) HPLC Analysis 96
3.6.2.4 High pH Anion Exchange Chromatography with Pulsed Amperometric Detection (HPAEC-PAD) 96
3.6.3 Capillary Electrophoresis (CE) 97
3.6.4 Fluorophore-assisted Carbohydrate Electrophoresis (FACE) and CGE-LIF 99
3.6.5 Mass Spectrometry (MS) 100
3.6.5.1 Ionization 100
3.6.5.2 Derivatization Techniques Used for MS Analysis of Glycans 102
3.6.5.3 Fragmentation of Carbohydrates 103
3.7 Conclusion 109
References 109
Part II Bioreactors 131
4 Bioreactors for StemCell and Mammalian Cell Cultivation 133
Ana Fernandes-Platzgummer, Sara M. Badenes, Cláudia L. da Silva, and JoaquimM. S. Cabral
4.1 Overview of (Mammalian and Stem) Cell Culture Engineering 133
4.1.1 Cell Products for Therapeutics 134
4.1.2 Cell as a Product: Stem Cells 136
4.2 Bioprocess Characterization 140
4.2.1 Cell Cultivation Methods 140
4.2.2 Cell Metabolism 141
4.2.3 Culture Medium Design 143
About the author
CLAIRE KOMIVES, PHD, is a Professor in the Chemical and Materials Engineering Department at San Jose State University. Her research interests focus on the development of low cost snake antivenom compounds.
WEICHANG ZHOU, PHD, is Chief Technology Officer, Biologics Development & Manufacturing at WuXi Biologics. He has previously worked at Genzyme Corporation and Merck Research Laboratories.
Summary
In the wake of rapid advances in genetic technologies, new products continue to be developed to help improve human health and quality of life.
