We recently finished our Ask the Expert discussion on Using Blood Cells As a Source for Generating Induced Pluripotent Stem Cells. This week we had many great questions and useful suggestions on issues including reprogramming methods, differentiation techniques, confirming pluripotency, and matrices. We also discussed advantages and challenges of utilizing blood cells over other cell sources.
This Ask the Expert Session was Sponsored by STEMCELL Technologies and hosted by Wing Chang, Scientist at STEMCELL Technologies. Wing Chang, PhD, joined STEMCELL Technologies’ Research and Development Department in 2012 as a Scientist and is responsible for the development of products for reprogramming and the generation of induced pluripotent stem cells. Prior to joining STEMCELL, Wing was a postdoctoral fellow at the Ottawa Hospital Research Institute and the University of Toronto.
While dermal fibroblasts were the first human cell type to be reprogrammed into iPS cells, blood cells are also increasingly being utilized as a starting cell type due to the limited invasiveness of sample collection, and the availability of banked blood samples representing a variety of disease, age, gender and geographical subtypes.
Check out this interesting discussion. Below is a sneak peek, for a full transcript, please see – Ask the Expert – Using Blood Cells As a Source for Generating Induced Pluripotent Stem Cells.
Can you speak to the advantages and challenges of using blood cells to generate iPSCs versus other cell types?
A couple of advantages of using blood cells as a source for generating iPSCs include the ease of obtaining blood samples from donors, and the higher reprogramming efficiency of blood cells compared to other cell types.
One of the challenges of using blood cells is determining which blood cell type to start with for iPSC generation and isolating/enriching for that population. This decision may be dependent on the amount of blood that you receive from the donor. If you use a heterogeneous population of mononuclear cells, you may generate iPSCs from progenitors as well as more mature cell types (e.g. T- and B-cells), the latter of which can give rise to can give rise iPSCs with genomic rearrangements. This is why we recommend using our Blood Reprogramming Kits for Erythroid Progenitor or CD34+ Progenitor Cells, as they include reagents to deplete T- and B-cells and enrich for only the desired starting cell type.
What are the differences in reprogramming cells on feeder cells vs feeder-free conditions?
Chan et al (1) demonstrated that reprogramming efficiency under feeder-free conditions was lower, but that all of the iPS cells generated were fully (not partially) reprogrammed. Therefore, it seems that culture conditions have an important role in the reprogramming process. That is why we recommend STEMCELL Technologies’ feeder-free, defined, and xeno-free media for reprogramming fibroblasts (TeSR™-E7™) or blood cells (ReproTeSR™), which generate recognizable iPS cells with less differentiated or partially reprogrammed background cell growth.
(1) Chan EM, et al. Nat Biotechnol 27(11): 1033–7, 2009
What do you recommend for differentiation of blood cell generated iPSCs – EBs or monolayer?
We typically perform directed differentiation of iPSCs (generated from blood cells) using monolayer cultures.
However, should you decide to differentiate using embryoid bodies (EBs), we would recommend using Aggrewell™ plates for reproducible production of uniformly-sized embryoid bodies.