Improved Transfection Efficiency and Metabolic Activity in Human Embryonic Stem Cell Using Non-Enzymatic Method

International Journal of Stem Cells
C-Yoon KimHyung-Min Chung

Abstract

Human embryonic stem cells (hESCs) are pluripotent cells widely used in conventional and regenerative medicine due to their ability to self-renew, proliferate and differentiate. Recently, genetic modification of stem cells using genome editing is the most advanced technique for treating hereditary diseases. Nevertheless, the low transfection efficiency of hESCs using enzymatic methods is still limited in in vitro preclinical research. To overcome these limitations, we have developed transfection methods using non-enzymatic treatments on hESCs. In this study, hESCs were transfected following enzymatic (TrypLE and trypsin) and non-enzymatic treatment ethylenediaminetetraacetic acid (EDTA) to increase transfection efficiency. Flow cytometric analysis using an enhanced green fluorescent protein vector showed a significantly increased transfection efficiency of EDTA method compared to standard enzyme method. In addition, the EDTA approach maintained stable cell viability and recovery rate of hESCs after transfection. Also, metabolic activity by using Extracellular Flux Analyzer revealed that EDTA method maintained as similar levels of cell functionality as normal group comparing with enzymatic groups. These results suggest that tran...Continue Reading

Methods Mentioned

BETA
genetic modification
transfection
transgenic
fluorescence activated cell sorting
FACS
PCR
fluorescence-activated cell sorting

Software Mentioned

BD CellQuest
SPSS Statistics
Extracellular Flux Analyzer

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