Issue 2, 2024

Evaluating Riboglow-FLIM probes for RNA sensing

Abstract

We recently developed Riboglow-FLIM, where we genetically tag and track RNA molecules in live cells through measuring the fluorescence lifetime of a small molecule probe that binds the RNA tag. Here, we systematically and quantitatively evaluated key elements of Riboglow-FLIM that may serve as the foundation for Riboglow-FLIM applications and further tool development efforts. Our investigation focused on measuring changes in fluorescence lifetime of representative Riboglow-FLIM probes with different linkers and fluorophores in different environments. In vitro measurements revealed distinct lifetime differences among the probe variants as a result of different linker designs and fluorophore selections. To expand on the platform's versatility, probes in a wide variety of mammalian cell types were examined using fluorescence lifetime imaging microscopy (FLIM), and possible effects on cell physiology were evaluated by metabolomics. The results demonstrated that variations in lifetime were dependent on both probe and cell type. Interestingly, distinct differences in lifetime values were observed between cell lines, while no overall change in cell health was measured. These findings underscore the importance of probe selection and cellular environment when employing Riboglow-FLIM for RNA detection, serving as a foundation for future tool development and applications across diverse fields and biological systems.

Graphical abstract: Evaluating Riboglow-FLIM probes for RNA sensing

Supplementary files

Transparent peer review

To support increased transparency, we offer authors the option to publish the peer review history alongside their article.

View this article’s peer review history

Article information

Article type
Communication
Submitted
12 Oct 2023
Accepted
03 Jan 2024
First published
08 Jan 2024
This article is Open Access
Creative Commons BY-NC license

RSC Chem. Biol., 2024,5, 109-116

Evaluating Riboglow-FLIM probes for RNA sensing

N. Sarfraz, L. K. Shafik, Z. R. Stickelman, U. Shankar, E. Moscoso and E. Braselmann, RSC Chem. Biol., 2024, 5, 109 DOI: 10.1039/D3CB00197K

This article is licensed under a Creative Commons Attribution-NonCommercial 3.0 Unported Licence. You can use material from this article in other publications, without requesting further permission from the RSC, provided that the correct acknowledgement is given and it is not used for commercial purposes.

To request permission to reproduce material from this article in a commercial publication, please go to the Copyright Clearance Center request page.

If you are an author contributing to an RSC publication, you do not need to request permission provided correct acknowledgement is given.

If you are the author of this article, you do not need to request permission to reproduce figures and diagrams provided correct acknowledgement is given. If you want to reproduce the whole article in a third-party commercial publication (excluding your thesis/dissertation for which permission is not required) please go to the Copyright Clearance Center request page.

Read more about how to correctly acknowledge RSC content.

Social activity

Spotlight

Advertisements