Biology subjects

Englert, D.

Publications and source records attributed to Englert, D..

3 recordsLinked to original sources

bioRxiv2022

Fast-exchanging spirocyclic rhodamine probes for aptamer-based super-resolution RNA imaging

Live-cell RNA imaging with high spatial and temporal resolution remains a major challenge. Here we report the development of RhoBAST:SpyRho, a fluorescent light-up aptamer (FLAP) system ideally suited for visualizing RNAs in live or fixed cells with various advanced fluorescence microscopy modalities. Overcoming problems associated with low cell permeability, brightness, fluorogenicity, and signal-to-background ratio of previous fluorophores, we designed a novel probe, SpyRho (Spirocyclic Rhodamine), which tightly binds to the RhoBAST aptamer. High brightness and fluorogenicity is achieved by shifting the equilibrium between spirolactam and quinoid. With its high affinity and fast ligand exchange, RhoBAST:SpyRho is a superb system for both super-resolution SMLM and STED imaging. Its excellent performance in SMLM and the first reported super-resolved STED images of specifically labeled RNA in live mammalian cells represent significant advances over other FLAPs. The versatility of RhoBAST:SpyRho is further demonstrated by imaging endogenous chromosomal loci and proteins.

biophysics↗

bioRxiv2021

Bright, fluorogenic and photostable avidity probes for RNA imaging

Fluorescent light-up aptamers (FLAPs) emerged as valuable tools to visualize RNA, but are mostly limited by poor brightness, low photostability and high fluorescence background. In this study, we combine bivalent (silicon) rhodamine fluorophores with dimeric FLAPs to yield bright and photostable complexes with low picomolar dissociation constants. Our avidity-based approach resulted in extreme binding strength and high fluorogenicity, enabling single mRNA tracking in living cells.

molecular biology↗

bioRxiv2020

RhoBAST - A Rhodamine-Binding Aptamer for Super-Resolution RNA Imaging

RhoBAST is a novel fluorescence light-up RNA aptamer (FLAP) that transiently binds a fluorogenic rhodamine dye. Fast dye association and dissociation result in intermittent fluorescence emission, facilitating single-molecule localization microscopy (SMLM) with an image resolution not limited by photobleaching. We demonstrate RhoBAST's excellent properties as a RNA marker by resolving subcellular and subnuclear structures of RNA in live and fixed cells by SMLM and structured illumination microscopy (SIM).

cell biology↗