Femtosecond laser assisted photo-transfection and differentiation of mouse embryonic stem cells

Lebogang Thobakgale; Sello Manoto; Satuurnin Ombinda Lemboumba; Malik Maaza; Patience Mthunzi-Kufa

doi:10.1117/12.2290206

13 February 2018 Femtosecond laser assisted photo-transfection and differentiation of mouse embryonic stem cells

Lebogang Thobakgale, Sello Manoto, Satuurnin Ombinda Lemboumba, Malik Maaza, Patience Mthunzi-Kufa

Proceedings Volume 10492, Optical Interactions with Tissue and Cells XXIX; 1049205 (2018) https://doi.org/10.1117/12.2290206
Event: SPIE BiOS, 2018, San Francisco, California, United States

Abstract

In tissue engineering research, stem cells have been used as starting material in the synthesis of mammalian cells for the treatment of various cell based diseases. This is done by manipulating the DNA content of the cells to induce a specific effect such as increased proliferation or developing a new cell type through the process of differentiation. Such controlled gene expression of stem cells is achieved by the method of transfection, where exogenous plasmid deoxyribonucleic acid (pDNA) is inserted into a stem cell using chemical, viral or physical methods. In this research, we used femtosecond (fs) laser pulses from a home-build microscope system to perforate the cellular membrane and allow entry of selected pDNA to alter the behaviour of mouse embryonic stem cells (mESCs). In one set of experiments, we induce fluorescence on mESCs using green fluorescence protein plasmid (pGFP) while in other tests; differentiation of mESCs into endoderm cells is performed using Sox-17 plasmid DNA (pSox-17). Primitive endoderm formation was thereafter confirmed using polymerase chain reactions (PCR) and the Sox-17 primer. Cell viability studies using adenosine triphosphate were also conducted. From the data, it was concluded that the photo-transfection method is biocompatible since it was able to induce fluorescence in mESCs. Secondly, it was confirmed that Sox-17 was photo-transfected successfully using 6 μW laser power, 128 fs pulses and 1kHz pulse repetition rate.

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Citation Download Citation

Lebogang Thobakgale, Sello Manoto, Satuurnin Ombinda Lemboumba, Malik Maaza, and Patience Mthunzi-Kufa "Femtosecond laser assisted photo-transfection and differentiation of mouse embryonic stem cells ", Proc. SPIE 10492, Optical Interactions with Tissue and Cells XXIX, 1049205 (13 February 2018); https://doi.org/10.1117/12.2290206

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