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Evaluating MINFLUX experimental performance in silico
Zach Marin1,2, Jonas Ries3,4,5
1Max Perutz Labs, Vienna Biocenter Campus (VBC), Vienna, Austria.
Nature Communications
|December 6, 2025
Summary
SimuFLUX, a new simulator, reveals performance limits in single-molecule localization microscopy (MINFLUX). It investigates how factors like background noise and fluorophore movement impact imaging resolution and tracking accuracy.
Area of Science:
- Biophysics
- Optical Microscopy
- Nanotechnology
Background:
- MINFLUX (MInimal reMoval oF foCUS) offers exceptional resolution for biological imaging and tracking.
- Idealized optical resolution of MINFLUX is known, but real-world performance is often degraded by system complexity and experimental imperfections.
Purpose of the Study:
- To develop a comprehensive and realistic simulator, SimuFLUX, for MINFLUX microscopy.
- To investigate the performance limitations of MINFLUX under various non-ideal conditions.
Main Methods:
- Development of SimuFLUX, a versatile simulation platform for MINFLUX.
- Utilizing the simulator to analyze the impact of fluorophore dynamics, background noise, localization algorithms, and optical misalignment on MINFLUX performance.
Main Results:
- Quantification of performance degradation due to specific experimental factors.
- Identification of key parameters influencing MINFLUX resolution and tracking fidelity.
- Demonstration of how simulation can guide experimental design and optimization.
Conclusions:
- SimuFLUX provides a powerful tool for understanding and overcoming MINFLUX limitations.
- Realistic simulation is crucial for optimizing MINFLUX experiments and achieving maximal resolution.
- The simulator aids in assessing the feasibility of complex MINFLUX applications.
