This past June I attended the International Society for Stem Cell Research (ISSCR) in Vancouver and over the next several weeks I will be sharing information that I gathered while I was there. The daily Innovation Showcases were particularity interesting and focused on new, state of the art products or approaches that enable stem cell culture and research. This first, in a series of blog posts, covers three of the Innovation Showcases.
Reprogramming Using Sendai Virus Technology
One interesting Innovation Showcase that I was able to attend was “Reprogramming Somatic Cells Using Sendai Virus Technology,” given by Dr. Laurence Daheron, Harvard Stem Cell Institute. The talk began with a discussion of the use of the Sendai Virus (SeV) for reporgramming Erythroblasts to iPSCs and the associated advantages of using the Sendai Virus method for reprogramming including the fact that it has been effective in a wide range of cell types.
While using SeV for reprogramming, Dr. Daheron’s lab attained good reprogramming efficiency and reliability, but faced challenges when it came to the speed in which the Sendai Virus was cleared and they needed faster elimination of the virus. Dr. Daheron presented data, which showed that after 6-8 passages, approximately 54% of the cells were still positive for Sendai Virus RNA and after 9-11 passages, approximately 21% were still positive. This clearance was not efficient enough for their application.
In November 2013, Life Technologies’ launched CytoTune 2.0, a second generation to their CytoTune iPS Sendai Reprogramming kit. By using CytoTune 2.0, Dr. Daheron’s lab was able to increase efficiency and achieve faster clearance of the Sendai Virus RNA. They found that with CytoTune 2.0 after 1-5 passages approximately 43% of the cells were positive, and after 6-8 passages only approximately 12.5% were positive. The ability to clear the Sendai Virus RNA more quickly enabled faster expansion and quicker time to experiments.
Other reported advantages of CytoTune 2.0 included:
- Requires only one overnight incubation
- Increased reprogramming efficiencies for more colonies
- Lower cytotoxicity to allow for smaller starting cell populations
- Easy to use
To view the entire presentation, please see – Reprogramming Somatic Cells Using Sendai Virus Technology
Animal Component-Free Culture System for Human Mesenchymal Stem Cells
I was also happy to be able to attend STEMCELL Technologies’ Innovation Showcase titled “A Novel Animal Component-Free Culture System for Efficient Derivation, Expansion and Cryopreservation of Human MSCs from Bone Marrow and Adipose Tissue.” The talk walked through their new line of animal-component free (ACF) products for human mesenchymal stem cells (MSCs) and touched on the reasons for developing a culture system that is free of animal components, including fetal bovine serum (FBS). There is concern about the use of FBS in the culture of stem cells, particularly in developing cell therapies due to the undefined nature of FBS and concerns about immune rejection of transplanted cells or disease transmission.
To address concerns about serum, STEMCELL Technologies has developed their MesenCult™-ACF line, which contains products for derivation, expansion, cryopreservation, and differentiation of human MSCs. To test these products, STEMCELL Technologies utilized protocols for both bone marrow- and adipose-derived MSCs and compared the performance of the ACF products to serum-containing medium.
I have selected some highlights from the data presented, but you can access the full presentation and all of the data here – A Novel Animal Component-Free Culture System for Efficient Derivation, Expansion and Cryopreservation of Human MSCs from Bone Marrow and Adipose Tissue
Derivation and Expansion:
MesenCult™-ACF Culture Kit and MesenCult™- ACF Dissociation Kit
According to data presented, in the bone marrow-derived protocol, MSCs in MesenCult™-ACF generated larger CFU-F colonies containing smaller cells and had higher expansion and faster doubling times than the serum control. There was also a reduction of hematopoietic cell contamination in early passages which resulted in a higher MSC yield and purity earlier than with the serum control. Similar results were found in the adipose tissue -derived protocol. In addition, data showed T-cell proliferation suppression with MesenCult™-ACF.
MesenCult™-ACF Freezing Medium
Data presented showed high recovery rates and differentiation potential was maintained.
MesenCult™-ACF Chondrogenic Differentiation Medium
MesenCult™ Adipogenic Differentiation Medium and MesenCult™ Osteogenic Differentiation Medium still contain serum, but animal component-free versions are in development.
Presented data demonstrating multi-lineage differentiation potential was maintained in vitro.
Single-Cell Biology – Powering Discovery
Fluidigm gave an interesting and informative presentation “Powering Discovery Through Single-Cell Biology: Unraveling Cell Fate, Differentiation and Lineage,” highlighting the importance of single-cell biology. In particular, due to the heterogeneous nature of stem and progenitor cells, using bulk analysis methods obscures the critical drivers of self-renewal, pluripotency, and differentiation occurring in individual cells. One way to address this challenge is to examine the cells individually, however, traditional methods such as FACS and laser dissection are not time or cost effective, require large sample inputs, and introduce experimental variability.
Fluidigm’s C1™Single-Cell Auto Prep System addresses these challenges by providing an automated workflow for isolating, staining, lysing and processing live viable cells using small sample amounts. In using the system for single-cell gene expression, miRNA analysis, or RNA or DNA Sequencing, 96 single-cells can be analyzed in under 24 hours.
Their newly launched C1 Open App Program allows researchers to develop custom scripts for the C1 system that meet specific research needs. This is particularly interesting as a tool to enable researchers to customize their workflows for their specific cell types and research goals. Later this year, Fluidigm is taking this approach one step further by creating a C1 Script Hub where researchers can share their custom scripts with others via an online hub. In turn, researchers will be able to search the hub for scripts that fit their needs. Hopefully the sharing of these research tools will help researchers avoid “reinventing the wheel” and will empower quicker development in new research fields.
To view additional information, please see Fluidigm’s C1™Single-Cell Auto Prep System
More to come..
This is just the first in a series of blogs covering ISSCR 2014. It was an excellent meeting and I highly recommend attending if you get the opportunity. Find out more about the ISSCR’s 2015 Annual Meeting in Stockholm, June 24-27 by visiting – ISSCR’s 2015 Annual Meeting