Impact of triplet state population on GFP-type fluorescence and photobleaching

IF 2.4 4区生物学 Q4 CELL BIOLOGY

Biology of the Cell Pub Date : 2025-02-13 DOI:10.1111/boc.202400076

Martin Byrdin, Svetlana Byrdina

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Abstract

Background Information

Based on recently published parameters (Rane et al. 2023, JPCB 127, 5046–5054) for (rs)EGFP triplet state formation and decay rates and yields, we consider the power density dependence of triplet state population dynamics and its consequences for the application of green fluorescent proteins in biological single molecule fluorescence microscopy.

Results

We find that under certain conditions, the photon budget of GFP type fluorescent proteins can be linearly dependent on power density and we propose a possible explanation for such a non-Hirschfeld photobleaching behavior. Moreover, illumination with millisecond pulses at sub-kHz rates is shown to improve photostability.

Conclusions

We stipulate that a judicious choice of excitation wavelength should take into account the triplet state absorption spectrum along with the singlet state absorption spectrum. Formulas are given for the estimation of the effects of such choice as function of the experimental parameters.

Significance

The linear photobleaching model as proposed by Hirschfeld 50 years ago with power-independent photon budget is not generally applicable to fluorescent proteins with millisecond-lived triplet states.

Abstract Image

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来源期刊

Biology of the Cell 生物-细胞生物学

CiteScore

5.30

自引率

0.00%

发文量

审稿时长

>12 weeks

期刊介绍： The journal publishes original research articles and reviews on all aspects of cellular, molecular and structural biology, developmental biology, cell physiology and evolution. It will publish articles or reviews contributing to the understanding of the elementary biochemical and biophysical principles of live matter organization from the molecular, cellular and tissues scales and organisms. This includes contributions directed towards understanding biochemical and biophysical mechanisms, structure-function relationships with respect to basic cell and tissue functions, development, development/evolution relationship, morphogenesis, stem cell biology, cell biology of disease, plant cell biology, as well as contributions directed toward understanding integrated processes at the organelles, cell and tissue levels. Contributions using approaches such as high resolution imaging, live imaging, quantitative cell biology and integrated biology; as well as those using innovative genetic and epigenetic technologies, ex-vivo tissue engineering, cellular, tissue and integrated functional analysis, and quantitative biology and modeling to demonstrate original biological principles are encouraged.