Correlation of mRNA delivery timing and protein expression in lipid-based transfection

A. Reiser; D. Woschée; N. Mehrotra; R. Krzyszton; H. H. Strey; J. O. Rädler

Recently, we established quantitative live-cell imaging on single-cell arrays (LISCA) for the time-resolved study of reporter gene expression at the single-cell level (14) (15) (16) . In case of mRNA-mediated eGFP expression, the single-cell fluorescence was found to follow a firstorder translation model, which renders the kinetics of mRNA expression with high predictive precision (15, 16) . Among others, the single-cell analysis infers the individual expression onset times after mRNA transfection. These findings open the door to establish a routine assay to assess the delivery timing in terms of a distribution of individual time-to-expression after transfection (17) . Furthermore, we automated single-cell fluorescence readout using microstructured substrates with arrays of single-cell adhesion sites combined with a tailored perfusion system enabling fluid exchange for mRNA transfection during the time-lapse measurement (18) . The platform provides access to the distribution of expression onsets and expression rates after transfection. In this context, a systematic study of nanocarrier uptake kinetics, the expression onset times, and how it relates to protein expression efficiency and external transfection conditions has not yet been carried out at the single-cell level.