Recombinant Protein Expression in Mammalian Cells: Methods and Protocols

✍ Scribed by David L. Hacker

• Publisher: Springer New York;Humana Press
• Year: 2018
• Tongue: English
• Leaves: 311
• Series: Methods in Molecular Biology 1850
• Edition: 1st ed.
• Category: Library

✦ Synopsis

This detailed volume explores advances in vector design, DNA delivery, cell cultivation, host cell engineering, and bioprocess optimization within the study of recombinant protein expression in mammalian cells. The majority of the protocols employ either Chinese hamster ovary cells (CHO) or human embryonic kidney 293 cells (HEK293), the workhorses of the field, as the production host; however, the methods can be adapted to other mammalian hosts under the appropriate cell-specific conditions. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.

Authoritative and convenient, Recombinant Protein Expression in Mammalian Cells: Methods and Protocols aims to aid researchers in building on our knowledge of protein structure and function and to speed the discovery of new therapeutic proteins.

✦ Table of Contents

Front Matter ....Pages i-xi
Transient Gene Expression in Suspension HEK293-EBNA1 Cells (Denis L’Abbé, Louis Bisson, Christian Gervais, Eric Grazzini, Yves Durocher)....Pages 1-16
Transient Expression of Recombinant Membrane-eGFP Fusion Proteins in HEK293 Cells (Joanna Pieprzyk, Samuel Pazicky, Christian Löw)....Pages 17-31
PEI-Mediated Transient Gene Expression in CHO Cells (Yashas Rajendra)....Pages 33-42
Stable Expression by Lentiviral Transduction of Cells (Natascha Gödecke, Hansjörg Hauser, Dagmar Wirth)....Pages 43-55
Inducible Protein Production in 293 Cells Using the piggyBac Transposon System (Iacovos P. Michael, Andras Nagy)....Pages 57-68
Recombinant CHO Cell Pool Generation Using piggyBac Transposon System (Sowmya Balasubramanian)....Pages 69-78
Genome Engineering of Hybridomas to Generate Stable Cell Lines for Antibody Expression (Cristina Parola, Derek M. Mason, Andreas Zingg, Daniel Neumeier, Sai T. Reddy)....Pages 79-111
Protein Expression via Transient Transfection of Mammalian Cells in a WAVE Bioreactor (David I. Fisher)....Pages 113-121
CHO and HEK293 Cultivation and Transfection in Single-Use Orbitally Shaken Bioreactors (David L. Hacker, Laurence Durrer, Soraya Quinche)....Pages 123-131
Bench-Scale Stirred-Tank Bioreactor for Recombinant Protein Production in Chinese Hamster Ovary (CHO) Cells in Suspension (Dominique T. Monteil, Jeffrey Kuan)....Pages 133-145
Continuous and Integrated Expression and Purification of Recombinant Antibodies (Sebastian Vogg, Moritz Kai Florian Wolf, Massimo Morbidelli)....Pages 147-178
High Throughput Transfection of HEK293 Cells for Transient Protein Production (Tia A. Arena, Peter D. Harms, Athena W. Wong)....Pages 179-187
Microfluidic Transfection for High-Throughput Mammalian Protein Expression (Kristina Woodruff, Sebastian J. Maerkl)....Pages 189-208
Genome-Wide High-Throughput RNAi Screening for Identification of Genes Involved in Protein Production (Sarah Inwood, Michael J. Betenbaugh, Madhu Lal, Joseph Shiloach)....Pages 209-219
Targeting miRNAs with CRISPR/Cas9 to Improve Recombinant Protein Production of CHO Cells (Kevin Kellner, Ankur Solanki, Thomas Amann, Nga Lao, Niall Barron)....Pages 221-235
Application of the CRISPR/Cas9 Gene Editing Method for Modulating Antibody Fucosylation in CHO Cells (Qiong Wang, Cheng-Yu Chung, Julian N. Rosenberg, Geng Yu, Michael J. Betenbaugh)....Pages 237-257
Scalable Production and Purification of Adeno-Associated Viral Vectors (AAV) (Daniel Blessing, Nicole Déglon, Bernard L. Schneider)....Pages 259-274
Considerations in the Use of Codon Optimization for Recombinant Protein Expression (Vincent P. Mauro, Stephen A. Chappell)....Pages 275-288
Versatile Cell-Free Protein Synthesis Systems Based on Chinese Hamster Ovary Cells (Lena Thoring, Stefan Kubick)....Pages 289-308
Back Matter ....Pages 309-311

✦ Subjects

Life Sciences; Protein Science