Single Molecule Localization via Phase-Modulated Structured Illumination
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Solution Overview
Problem
Current methods for measuring the central location of a single molecule are limited by accuracy due to the division of photons among pixels and sensitivity/noise differences, and are restricted to small areas with concentrated light patterns.
Innovation Solution
A device and method using structured illumination and phase detection, where laser light is formed into a periodic pattern, and the phase is modulated to measure the central location of a single molecule based on fluorescence emission intensity across multiple images, allowing for improved accuracy and measurement of molecules in arbitrary locations and structures across large areas.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If a pixelated two-dimensional image is approximated with a two-dimensional function (Gaussian or Airy function) to measure central location, then measurement can be performed with general fluorescent molecules, but the accuracy is limited to several tens of nanometer due to photon division among pixels and sensitivity/noise differences
Solution Approach 1:
The patent segments the measurement process into multiple phase steps, acquiring images at different phase positions (0, 120, 240 degrees) of the structured illumination pattern. This segmentation allows the system to extract phase information from multiple measurements, improving the precision of central location determination while efficiently utilizing the limited photon budget by distributing measurements across phases rather than relying on a single image.
2Measurement precision
If donut shaped light with central intensity of 0 is used to excite fluorescent molecules, then accuracy of several nanometers can be obtained with smaller number of photons, but only the location of molecules confined in a small area can be measured
Solution Approach 1:
The patent employs a dynamic structured illumination pattern that can be phase-modulated across different positions. By systematically varying the phase of the periodic illumination pattern (0, 120, 240 degrees) and scanning across the sample area, the system extends the measurable area beyond the confined region of static donut-shaped light while maintaining high measurement precision through phase detection analysis.
3Measurement precision
If structured illumination with periodic pattern and phase modulation is used to measure central location, then accuracy is improved and locations of molecules distributed in arbitrary location with arbitrary structure in large area can be measured, but device complexity increases
Solution Approach 1:
The patent utilizes periodic structured illumination patterns with well-defined phase relationships (0, 120, 240 degrees). This periodic action simplifies the control complexity by using regular, repeating patterns that can be generated using standard optical components such as diffraction gratings or spatial light modulators. The phase-modulated periodic illumination enables precise measurements while maintaining manageable device complexity through the use of periodic rather than arbitrary illumination sequences.
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 enhances the accuracy of single molecule location measurement with low photons, improves image resolution, and enables ultra-high resolution optical imaging of arbitrary-sized and shaped structures by scanning periodic patterns across a large area, surpassing the limitations of existing technologies.
Implementation Method 1
forms laser light into light having a predetermined periodic pattern to generate a structured illumination
Implementation Method 2
detect single molecule signal which is generated from the target object to be generated by the structured illumination
Data Source
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AI summary
According to the present disclosure, an optical signal emitted from a single molecule is received to measure a central location of the single molecule while changing a phase of a structured illumination having a periodic pattern to measure a phase of a pattern in which a fluorescence intensity is periodically changed in accordance with a distance between the pattern and the single molecule while displacing the periodic pattern by a specific interval to measure the central location of the single molecule, thereby improving an accuracy of the central location of the single molecule with low photons and as a result, the resolution of the image may be enhanced.