Molecule Localization via Zero-Point Excitation Intensity
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Solution Overview
Problem
Current methods for determining the position of singularized molecules excitable with excitation light for luminescence emission face challenges such as bleaching and limited precision due to high photon requirements and directional light distribution, which hinder accurate tracking and localization beyond the diffraction barrier.
Innovation Solution
A method involving directing excitation light with a zero-point intensity distribution onto the sample, registering luminescence light at different zero-point positions, and deducing the molecule's position from these intensities, allowing for spatial high-resolution localization without excessive photon emission and bleaching.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If a plurality of photons of the luminescence light is detected to determine the position of a molecule with precision below the diffraction barrier, then the measurement precision is improved, but the molecule is bleached during or prior to determining its position
Solution Approach 1:
The patent segments the measurement process into multiple frames, where only a sparse subset of molecules is activated in each frame. This allows position determination with sufficient precision while limiting the total photons each molecule emits before bleaching, thus resolving the contradiction between measurement precision and molecule stability
Solution Approach 2:
The patent employs periodic activation of molecules in a sparse subset manner across different frames. Molecules are activated periodically rather than continuously, allowing position determination while preventing excessive photon emission that would cause bleaching, thereby maintaining both precision and reliability
2Illumination intensity
If excitation light with high intensity is used to excite molecules for luminescence emission, then the luminescence signal is strengthened, but the molecules undergo bleaching
Solution Approach 1:
The patent applies local quality by activating only a sparse subset of molecules in each frame rather than all molecules. This localized activation ensures sufficient luminescence signal from active molecules while preventing bleaching of inactive molecules, resolving the contradiction between signal intensity and molecule stability
Solution Approach 2:
The patent uses periodic activation where molecules are excited in a time-varying sparse subset pattern across frames. This periodic excitation provides strong luminescence signals when molecules are active while limiting cumulative exposure to prevent bleaching, balancing signal intensity and reliability
3Measurement precision
If the distance between singularized molecules is kept at least as high as the diffraction barrier, then the luminescence light from different molecules can be registered separately, but the spatial resolution is limited to above the diffraction barrier
Solution Approach 1:
The patent applies preliminary action by determining positions of molecules in a time-varying sparse subset pattern before they can interfere with each other. This allows super-resolution localization below the diffraction barrier by capturing positional information when molecules are spatially separated in time, resolving the contradiction between spatial resolution and minimum distance
Solution Approach 2:
The patent uses periodic activation of molecular subsets to achieve super-resolution. By periodically activating sparse subsets and determining positions when molecules are naturally separated, the system achieves spatial resolution below the diffraction barrier without requiring fixed large distances between all molecules, balancing measurement precision and spatial constraints
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
This approach achieves spatial resolution below the diffraction barrier, enabling precise localization of singularized molecules with reduced photon usage and minimizing bleaching, thereby improving tracking and imaging capabilities.
Implementation Method 1
a singularized molecule which is excitable with excitation light for emission of luminescence light
Implementation Method 2
the position of the molecule can be determined from the spatial distribution of these photons over the detector
Data Source
AI summary
For spatial high resolution determining a position of a singularized molecule, which is excitable with excitation light for emission of luminescence light, in n spatial dimensions in a sample, the excitation light is directed onto the sample with an intensity distribution, which has a zero point and intensity increasing regions adjoining the zero point on both sides in each of the n spatial dimensions. The zero point is arranged at not more than n×3 different positions. The luminescence light emitted by the singularized molecule is separately registering for each of the different positions of the zero point. The position of the singularized molecule in the n spatial dimensions in the sample is deduced from intensities of the luminescence light separately registered for the not more than n×3 different positions of the zero point.