Minflux

Minflux microscopy, often shortened to Minflux, is a single-molecule localization microscopy technique that improves upon super-resolution microscopy methods like stimulated emission depletion (STED) microscopy and photoactivated localization microscopy (PALM)/stochastic optical reconstruction microscopy (STORM). It achieves superior spatial resolution and temporal resolution compared to these earlier techniques.

The key principle of Minflux involves iteratively minimizing the excitation volume around a fluorophore until the fluorophore signal is maximized. This is typically accomplished by using a doughnut-shaped excitation beam. The position of the fluorophore is then determined with high precision based on the known characteristics of the excitation beam and the detected fluorescence signal. The method relies on a feedback loop to repeatedly refine the excitation position, effectively 'tracking' and precisely localizing the fluorophore.

Several variants of Minflux exist, each with specific implementations and advantages. These variations can differ in the excitation patterns used (e.g., using a doughnut-shaped beam, a Gaussian beam with rapid scanning, or pulsed interleaved excitation) and the computational methods employed for localization.

Minflux microscopy offers several advantages over other super-resolution techniques:

• High Spatial Resolution: Minflux can achieve resolutions down to a few nanometers, allowing for detailed visualization of cellular structures.
• High Temporal Resolution: The rapid localization capabilities of Minflux enable tracking of dynamic processes in live cells with high temporal resolution.
• Low Photobleaching: The efficient excitation scheme of Minflux minimizes the amount of light required to localize fluorophores, reducing photobleaching and allowing for longer observation times.
• Compatibility with various fluorophores: Minflux can be adapted to work with a range of fluorescent dyes and proteins.

However, Minflux also presents some challenges:

• Instrumentation Complexity: Minflux requires sophisticated instrumentation and control systems, which can be expensive and complex to operate.
• Data Analysis: The data generated by Minflux experiments requires specialized analysis techniques to extract accurate localization information.
• Sample Preparation: Similar to other super-resolution techniques, sample preparation is critical for achieving optimal results.

Minflux is applied in diverse fields, including cell biology, neurobiology, and materials science, to study molecular structures and dynamics at the nanoscale. It has been used to investigate protein interactions, receptor dynamics, and the organization of cellular compartments.