Fast, Efficient, and Gentle Transfection of Human Adherent Cells in Suspension.

We demonstrate a highly efficient method for gene delivery into clinically relevant human cell types, such as induced pluripotent stem cells (iPSCs) and fibroblasts, reducing the protocol time by one full day. To preserve cell physiology during gene transfer, we designed a microfluidic strategy, which facilitates significant gene delivery in a short transfection time (<1 min) for several human cell types. This fast, optimized and generally applicable cell transfection method can be used for rapid screening of different delivery systems and has significant potential for high-throughput cell therapy applications.

[1]  V. Adami,et al.  An electroporation protocol for efficient DNA transfection in PC12 cells , 2013, Cytotechnology.

[2]  R J Roselli,et al.  A model for the analysis of nonviral gene therapy , 2003, Gene Therapy.

[3]  T. Reineke,et al.  Poly(glycoamidoamine)s for Gene Delivery: Stability of Polyplexes and Efficacy with Cardiomyoblast Cells , 2006 .

[4]  James M Piret,et al.  Mathematical model of the rate‐limiting steps for retrovirus‐mediated gene transfer into mammalian cells , 2010, Biotechnology and bioengineering.

[5]  D. Di Carlo Inertial microfluidics. , 2009, Lab on a chip.

[6]  J. Wagner,et al.  Induced pluripotent stem cells from individuals with recessive dystrophic epidermolysis bullosa. , 2011, The Journal of investigative dermatology.

[7]  T. Reineke,et al.  Membrane and nuclear permeabilization by polymeric pDNA vehicles: efficient method for gene delivery or mechanism of cytotoxicity? , 2012, Molecular Pharmaceutics.

[8]  T. Reineke,et al.  Exploring the mechanism of plasmid DNA nuclear internalization with polymer-based vehicles. , 2012, Molecular pharmaceutics.

[9]  Yong Jin,et al.  Slurry Reactors for Gas-to-Liquid Processes: A Review , 2007 .

[10]  Peter W. Zandstra,et al.  Derivation, expansion and differentiation of induced pluripotent stem cells in continuous suspension cultures , 2012, Nature Methods.

[11]  S. Orkin,et al.  Reprogramming Committed Murine Blood Cells to Induced Hematopoietic Stem Cells with Defined Factors , 2014, Cell.

[12]  Chih-Kuang Chen,et al.  Overcoming nonviral gene delivery barriers: perspective and future. , 2013, Molecular pharmaceutics.

[13]  B. Palsson,et al.  Retroviral infection is limited by Brownian motion. , 1996, Human gene therapy.

[14]  D. Prescott,et al.  An evaluation of the double thymidine block for synchronizing mammalian cells at the G1-S border. , 1971, Experimental cell research.

[15]  N. Ingle,et al.  Polymeric nucleic acid vehicles exploit active interorganelle trafficking mechanisms. , 2013, ACS nano.

[16]  Daniel G. Anderson,et al.  Non-viral vectors for gene-based therapy , 2014, Nature Reviews Genetics.

[17]  H. Harashima,et al.  Endocytosis of gene delivery vectors: from clathrin-dependent to lipid raft-mediated endocytosis. , 2013, Molecular Therapy.

[18]  Jeffry D. Sander,et al.  CRISPR-Cas systems for editing, regulating and targeting genomes , 2014, Nature Biotechnology.