iPSC generation - Human iPSC colonies expressing pluripotency markers

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L7™ PBMC Reprogramming Bundle

For robust generation of iPSCs from peripheral blood monunuclear cells (PBMCs).

iPSC Generation 

For iPSC generation, various adult cell types including peripheral blood mononuclear cells (PBMCs), fibroblasts or CD34+ hematopoietic progenitor cells can be reprogrammed to a pluripotent state by introduction of defined transcription factors.  The resulting induced pluripotent stem cells (iPSCs) appear to be phenotypically and functionally equivalent to human embryonic stem cells (hESCs). The ability of hESCs and iPSCs to self-renew in culture and their potential to differentiate into cell types from all three germ layers makes them a powerful research tool.  

iPSC generation and potential applications:

  • Identifying the mechanisms of human development
  • Generating new models to study human disease pathogenesis
  • Production of specific cell types for in vitro drug screening and toxicity studies  

Moreover, iPSCs also have great potential as a renewable source of material for producing therapeutic cells for transplantation in regenerative medicine. Because they can be derived from any individual, iPSCs may be useful for generating autologous therapeutic cells for transplantation.  


Nucleofector™ Technology for iPSC Generation

For human cells, this cellular reprogramming process was first described in 2007 by two independent research groups (Yu J et al. (2007) Science 318 (5858): 1917–1920; Takahashi K et al. (2007) Cell 131 (5): 861–872). These reprogramming factors were initially transferred into the somatic cells by retroviral transduction, thereby diminishing their applicability to the clinic. Lonza’s Nucleofector™ Technology has been demonstrated to be a convenient, efficient, and cost-effective non-viral alternative for iPSC generation and is currently being used by leading scientists around the world.

Benefits of the Nucleofector™ Technology for iPSC generation include:

  • Simple, single-step procedure to introduce DNA/RNA, e.g. episomal vectors
  • Successfully tested for generation of iPSCs from various cell types
  • Seamless combination with Lonza’s L7™ hPSC Culture System for feeder- and xeno-free iPSC culture
  • Availability of 4D-Nucleofector™ Kits with GMP Solutions
  • Also proven for efficient iPSC transfection, e.g. for genome editing using ZFN, TALEN, and CRISPR systems

 

Guidelines for iPSC Generation from Various Starting Cell Types

a) Human PBMCs

For reprogramming of peripheral blood mononuclear cells (PBMCs), Lonza’s Pluripotent Stem Cell Innovation Team has developed an optimized protocol using the 4D-Nucleofector™ System and episomal vectors. The protocol is accompanied by supporting products, i.e. the L7™ PBMC Reprogramming Bundle

Positive control cells:   Human Peripheral Blood Mononuclear Cells
Cell-specific media:   L7™ PBMC Priming-Recovery Kit 
4D-Nucleofector™ Kit:   P3 Primary Cell 4D-Nucleofector™ Kit 
iPSC Culture:   L7™ hPSC Culture System 
References or protocols:  
  • Reprogramming protocol - Available soon!
  • Tran HT et al. (2014) – A Case Study (white paper)

   

b) Human fibroblasts

For reprogramming of human dermal fibroblasts, Lonza does not have a specific reprogramming protocol yet, but there are various references available that can provide you with some details.

Positive control cells:    Human Dermal Fibroblasts 
Cell-specific media:    FGM™ Medium 
4D-Nucleofector™ Kit:    P2 Primary Cell 4D-Nucleofector™ Kit  
iPSC Culture:    L7™ hPSC Culture System 
References or protocols:   

 

c) Human CD34+ hematopoietic progenitors

For reprogramming of human CD34+ hematopoietic progenitors, Lonza does not have a specific reprogramming protocol yet, but there are various references available that can provide you with some details.

Positive control cells:   

Human Bone Marrow CD34+ Cells

Human Cord Blood CD34+ Cells

Cell-specific media:   
4D-Nucleofector™ Kit:    P3 Primary Cell 4D-Nucleofector™ Kit 
iPSC Culture:    L7™ hPSC Culture System 
References or protocols:   

 

d) Other cell types

Besides the three main cell types used as starting material for iPSC generation, the Nucleofector™ Technology has also been used to reprogram various other cell types.

Positive control cells:   

Human Keratinocytes

Human Bone Marrow Mononuclear Cells

Human Neural Progenitor Cells 

Human Adipose-derived Stem Cells

Cell-specific media:    See respective cell type or reference 
4D-Nucleofector™ Kit:    See Nucleofector™ Kits for Primary Cells 
iPSC Culture   L7™ hPSC Culture System 
References or protocols:   

 

iPSC generation does not have to be a mystery with Lonza’s comprehensive solutions - From the most commonly used primary cell types for reprogramming; to the Nucleofector™ Technology for efficient, non-viral delivery of reprogramming factors; to the L7™ Culture System for feeder-free, xeno-free culture of human iPSCs, Lonza provides a wide range of products to aid in your generation and culturing of iPSCs.