Fluorescence Correlation Spectroscopy to find the critical balance between extracellular association and intracellular dissociation of mRNA complexes

Acta Biomaterialia
Heyang ZhangK Remaut

Abstract

Fluorescence Correlation Spectroscopy (FCS) is a promising tool to study interactions on a single molecule level. The diffusion of fluorescent molecules in and out of the excitation volume of a confocal microscope leads to the fluorescence fluctuations that give information on the average number of fluorescent molecules present in the excitation volume and their diffusion coefficients. In this context, we complexed mRNA into lipoplexes and polyplexes and explored the association/dissociation degree of complexes by using gel electrophoresis and FCS. FCS enabled us to measure the association and dissociation degree of mRNA-based complexes both in buffer and protein-rich biological fluids such as human serum and ascitic fluid, which is a clear advantage over gel electrophoresis that was only applicable in protein-free buffer solutions. Furthermore, following the complex stability in buffer and biological fluids by FCS assisted to understand how complex characteristics, such as charge ratio and strength of mRNA binding, correlated to the transfection efficiency. We found that linear polyethyleneimine prevented efficient translation of mRNA, most likely due to a too strong mRNA binding, whereas the lipid based carrier Lipofectamine®...Continue Reading

Citations

Jun 5, 2019·Analytical and Bioanalytical Chemistry·Di SuJicun Ren
May 6, 2020·Nanoscale·Lies A L FliervoetTina Vermonden
Sep 13, 2020·International Journal of Molecular Sciences·Shuqin XuLu Zhang
Feb 6, 2021·Pharmaceutics·Basmah N AldosariAlanood S Almurshedi
Apr 11, 2021·Journal of Controlled Release : Official Journal of the Controlled Release Society·Annalisa RossoGiovanna Lollo
May 4, 2021·Chemical Reviews·Ramya KumarTheresa M Reineke
Jul 3, 2021·Pharmaceutics·Christophe DeleheddeFederico Perche
Jun 17, 2021·Acta Biomaterialia·Minsong GaoJianzhao Liu
Aug 21, 2021·International Journal of Pharmaceutics·Qi ShuaiYunfeng Yan

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