Light Microscope Emitter Localization With MINFLUX Error Correction
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Solution Overview
Problem
Existing MINFLUX localization methods suffer from suboptimal photon efficiency and position accuracy due to systematic deviations between initial position estimates and final determinations, leading to inefficient use of photons and reduced localization precision.
Innovation Solution
A two-step localization method that includes an initial localization step followed by a MINFLUX-based step with systematic deviation correction, using illumination positions arranged around the emitter's location to enhance accuracy, and employing correction values or vectors to align the localization steps.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If a single-step MINFLUX localization method is used, then the method is simple and fast, but systematic deviations between initial position estimates and final determinations reduce position accuracy and photon efficiency
Solution Approach 1:
The localization process is divided into two distinct steps: a first localization step that provides an initial position estimate, and a second localization step that refines the position using MINFLUX illumination patterns. This segmentation allows each step to optimize for its specific function, with the first step prioritizing speed and the second step prioritizing accuracy, thereby resolving the contradiction between productivity and measurement precision.
Solution Approach 2:
The first localization step performs a preliminary localization to obtain an initial position estimate before the more complex second MINFLUX localization step. This preliminary action provides a starting point that reduces the computational and photonic burden of the second step, enabling high accuracy without sacrificing the overall speed of the localization process.
2Measurement precision
If correction values are applied to compensate for systematic deviations, then position accuracy improves, but device complexity and computational requirements increase
Solution Approach 1:
The method incorporates feedback by using the results of the first localization step to inform and correct the second localization step. Correction values or vectors are applied based on the systematic deviations observed, creating a feedback loop that continuously refines position accuracy without requiring complete redesign of the localization system.
Solution Approach 2:
The method adjusts localization parameters by introducing correction values or vectors that modify the position determination based on systematic deviations. These parameter changes are applied selectively in the second localization step, improving accuracy without fundamentally changing the overall method structure or increasing excessive complexity.
3Measurement precision
If illumination light is applied to achieve high position accuracy, then localization precision improves, but photon efficiency decreases due to systematic deviations causing inefficient photon use
Solution Approach 1:
The illumination process is segmented into two phases: the first localization step uses minimal illumination to obtain an initial estimate, and the second step uses targeted MINFLUX illumination patterns only where needed. This segmentation prevents unnecessary photon exposure and optimizes photon efficiency while maintaining high localization precision.
Solution Approach 2:
The preliminary first localization step provides an initial position estimate that guides the second illumination step, ensuring that photons are applied only where necessary to refine the position. This preliminary action prevents inefficient photon use by avoiding redundant illumination and focusing resources on achieving the required precision.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
Improves photon efficiency and position accuracy by minimizing systematic errors, allowing precise localization of individual emitters with reduced light exposure, overcoming limitations of prior methods.
Implementation Method 1
The individual emitters are in particular fluorophores and the illumination light is in particular excitation light, which excites the fluorophores, whereupon these emit fluorescent light
Implementation Method 2
The light that induces or modulates the light emission of the particles may also be STED (stimulated emission depletion) light
Data Source
AI summary
A method for localizing individual emitters in a sample with a first localization step and a second localization step with an increased accuracy including illuminating the emitter with an intensity distribution of the illumination light having a local minimum at illumination positions arranged around the location of the emitter determined in the first localization step, detecting the light emissions of the emitter for the illumination positions, and determining the location of the emitter from the light emissions detected for the illumination positions, wherein a location correction compensating for the systematic deviation is applied to determine the illumination positions in the second localization step, as well as a light microscope and a computer program for carrying out the method.