MINFLUX
Minimal Photon Fluxes Microscopy
Services offered
SMLM
Single Molecule Localisation Microscopy
STED
Stimulated Emission Depletion Microscopy
Learn more about MINFLUX
The imaging of minimal photon fluxes combines a coordinate-targeted single point scanning approach with the localization of stochastically activated single fluorophores to provide the currently highest resolving form of super-resolution microscopy.
Upon encountering a single point source signal, the exact position of a single fluorophore is interpolated through the photons collected at different positions of a series of patterns with decreasing search ranges that are executed by the probing beam (a concentric illumination ring with a central minimum) to home in on the fluorophore position. The combination of beam targeting with single molecule detection thus allows a localization accuracy in the nanometer range while only requiring a minimal number of photon detections overall. The MINFLUX principle can be applied for highly resolved imaging (stochastically activated fluorophores in a sample are imaged sequentially and integrated into a set of coordinates) as well as for the dynamic tracing of a single fluorescent molecule at unprecedented temporal and spatial resolution.
Learn more about SMLM
In SMLM is based on the fact that in diffraction-limited optics the position of a single point (i.e. a single molecule) can be determined with much higher accuracy than the minimal resolvable distance between two points.
Applying a fit for the central maximum of a single point source can thus determine its position with an accuracy that can be an order of magnitude higher than the optical resolution of the imaging system used. This cannot be applied in standard imaging as single point sources cannot be separated from their neighbours while all of them are detected together. In SMLM the label is made sufficiently scarce for the detection of single molecule positions by either the sparse photoconversion of single molecules to an active state (Photoactivation Localisation Microscopy, PALM), by the induction of transient darkstates in a vast majority of the fluorophores in the image field with only randomly distributed single emitters on at any time (Stochastic Optical Reconstruction Microscopy, STORM) or by the transient binding of diluted fluorescent tags to their targets (Points Accumulation for Imaging in Nanoscale Topography, PAINT).
Learn more about STED
In STED microscopy, an excited fluorescent molecule is induced to return to the ground state through the interaction with a photon of a wavelength inside the fluorophore’s emission spectrum.
This process is called stimulated emission. The combination of a focused excitation beam with a phase-shifted depletion beam in the same position creates a central excitation volume that is surrounded by a concentric ring of depletion light that deactivates fluorophores in its area and thus will reduce the effective fluorescence detection volume. In combination with confocal scanning this allows imaging below the diffraction limit. The resolution improvement is determined by the intensity of the applied depletion and can either be applied directly or be further enhanced by analysing the distribution of shortened fluorescence lifetimes that are caused by the interaction with the depletion beam.
