1. Successful Generation of Human Induced Pluripotent Stem Cell Lines from Blood Samples Held at Room Temperature for up to 48 hr
- Author
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Alex Alderton, Filipa A.C. Soares, Christopher M. Kirton, Chukwuma A. Agu, Minal Patel, Jonathan Lineham, Ludovic Vallier, Sharad R. Patel, Sally Forrest, Fengtang Yang, and Rizwan Ansari
- Subjects
Resource ,Cellular differentiation ,Induced Pluripotent Stem Cells ,Germ layer ,Cell Separation ,Biology ,Biochemistry ,Peripheral blood mononuclear cell ,Cryopreservation ,Genomic Instability ,03 medical and health sciences ,0302 clinical medicine ,Genetics ,Humans ,Cellular Reprogramming Techniques ,Induced pluripotent stem cell ,lcsh:QH301-705.5 ,Cells, Cultured ,030304 developmental biology ,0303 health sciences ,lcsh:R5-920 ,Temperature ,Cell Differentiation ,Cell Biology ,Cellular Reprogramming ,Sample stability ,Cell biology ,lcsh:Biology (General) ,Blood Preservation ,Leukocytes, Mononuclear ,lcsh:Medicine (General) ,Reprogramming ,030217 neurology & neurosurgery ,Developmental Biology - Abstract
Summary The collection sites of human primary tissue samples and the receiving laboratories, where the human induced pluripotent stem cells (hIPSCs) are derived, are often not on the same site. Thus, the stability of samples prior to derivation constrains the distance between the collection site and the receiving laboratory. To investigate sample stability, we collected blood and held it at room temperature for 5, 24, or 48 hr before isolating peripheral blood mononuclear cells (PBMCs) and reprogramming into IPSCs. Additionally, PBMC samples at 5- and 48-hr time points were frozen in liquid nitrogen for 4 months and reprogrammed into IPSCs. hIPSC lines derived from all time points were pluripotent, displayed no marked difference in chromosomal aberration rates, and differentiated into three germ layers. Reprogramming efficiency at 24- and 48-hr time points was 3- and 10-fold lower, respectively, than at 5 hr; the freeze-thaw process of PBMCs resulted in no obvious change in reprogramming efficiency., Graphical Abstract, Highlights • hIPSCs were derived from blood samples stored for 2 days at room temperature • Blood held for 24 and 48 hr produced less IPSC colonies than at 5 hr • Increasing input cells and virus volumes rescued the drop in reprogramming efficiency • IPSCs from all three storage time points can differentiate into three germ layers, Agu and colleagues investigated the permissible time during which blood samples can be stored at room temperature after collection without severely compromising their ability to be successfully reprogrammed into hIPSCs. They found that IPSCs can be derived from blood stored for 2 days at room temperature. The derived IPSCs maintained genomic stability and can differentiate into three germ layers.
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