顕微鏡 従来の分解能限界を克服し、分子の高速共同追跡を実現(Microscopy: Overcoming the traditional resolution limit for the fast co-tracking of molecules)

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2024-02-12 ミュンヘン大学(LMU)

LMUの研究チームは、pMINFLUX多重化という新しい超解像顕微鏡法を開発し、Nature Photonics誌に発表しました。この手法は、染料の点滅を利用せずに複数の生体分子の位置を同時に特定することができ、1ナノメートルの精度で位置を特定し、蛍光寿命を測定することができます。この技術は、タンパク質の相互作用などの生物学的現象の新たな洞察を提供する可能性があります。

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pMINFLUXにおける超分解能FRETと共同追跡 Super-resolved FRET and co-tracking in pMINFLUX

Fiona Cole,Jonas Zähringer,Johann Bohlen,Tim Schröder,Florian Steiner,Martina Pfeiffer,Patrick Schüler,Fernando D. Stefani & Philip Tinnefeld
Nature Photonics  Published:09 February 2024
DOI:https://doi.org/10.1038/s41566-024-01384-4

顕微鏡 従来の分解能限界を克服し、分子の高速共同追跡を実現(Microscopy: Overcoming the traditional resolution limit for the fast co-tracking of molecules)

Abstract

Single-molecule fluorescence resonance energy transfer (smFRET) is widely used to investigate dynamic (bio)molecular interactions occurring over distances of up to 10 nm. Recent advances in super-resolution methods have brought their spatiotemporal resolution closer towards the smFRET regime. Although these methods do not suffer from the spatial restrictions of FRET, they only visualize one emitter at a time, thus making it difficult to capture fast dynamics of the interactions. Here we describe two approaches to overcome this limitation in pulsed-interleaved MINFLUX (pMINFLUX) microscopy by using its intrinsic fluorescence lifetime information. First we combine pMINFLUX with smFRET, which enables tracking a FRET donor with nanometre precision while simultaneously determining its distance to a FRET acceptor, yielding the acceptor position by multilateration. Second, we developed pMINFLUX lifetime multiplexing—a method that simultaneously tracks two fluorophores with similar spectral properties but distinct fluorescence lifetimes—to extend co-localized tracking beyond the FRET range. We demonstrate applications on DNA origami systems as well as by imaging the paratopes of an antibody with precision better than 2 nm, paving the way for nanometre precise co-localized tracking for inter-dye distances between 4 nm and 100 nm, and closing the resolution gap between smFRET and co-tracking.

1700応用理学一般
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