Zero-Mode Waveguide Arrays for Single-Molecule Sequencing
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Solution Overview
Problem
Conventional DNA sequencing methods, such as dideoxy sequencing, are complex and costly due to the need for sample preparation to ensure identical DNA molecules in multiple tubes, and single-molecule analysis techniques like NSOM and SMS require low concentrations, limiting the study of concentration-dependent molecule dynamics.
Innovation Solution
A high-sensitivity and high-throughput system for analyzing multiple molecules by detecting light emissions from single molecules using a sample holder with spatially separated source points, a light source, and an optical assembly to collect and analyze fluorescent signals, allowing for simultaneous sequencing and SNP detection.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional dideoxy sequencing is used, then DNA sequencing can be performed, but sample preparation complexity and cost increase
Solution Approach 1:
The invention divides the DNA sequencing process into individual single-molecule observations rather than bulk population analysis. Each DNA molecule is sequenced independently in separate observation volumes, eliminating the need for complex sample preparation to ensure identical molecules across multiple tubes. This segmentation approach maintains sequencing accuracy while dramatically simplifying sample preparation.
Solution Approach 2:
The invention introduces zero-mode waveguides as intermediary structures that confine and isolate single DNA molecules in zeptoliter-scale observation volumes. These waveguides act as mediators between the DNA molecules and detection systems, enabling single-molecule observation without requiring complex sample preparation or low concentration conditions.
2Measurement precision
If NSOM and SMS techniques are used for single molecule analysis, then molecular level investigation capability is improved, but sample concentration must be reduced to picomolar to nanomolar levels
Solution Approach 1:
The invention fundamentally changes the observation volume parameter from femtoliter-scale (NSOM/SMS) to zeptoliter-scale using zero-mode waveguides. This parameter change enables single-molecule detection at natural physiological concentrations rather than requiring dilution to picomolar or nanomolar levels, thereby maintaining both detection sensitivity and physiological relevance.
3Quantity of substance
If zero-mode waveguides are used for single molecule analysis, then observation volume is reduced to zeptoliter scale, but device complexity and manufacturing difficulty increase
Solution Approach 1:
The invention creates arrays of zero-mode waveguide structures that can be manufactured using standard semiconductor fabrication techniques. By copying and replicating the waveguide geometry across the substrate, the system achieves multiple observation volumes while maintaining ease of manufacture through established fabrication processes rather than requiring complex custom fabrication for each observation volume.
4Productivity
If multiple DNA molecules are analyzed simultaneously in bulk, then throughput is improved, but ability to study concentration-dependent molecule dynamics is lost
Solution Approach 1:
The invention segments the analysis into parallel single-molecule observations in multiple zero-mode waveguides. Each waveguide contains and analyzes individual DNA molecules independently, enabling simultaneous high-throughput sequencing while maintaining the ability to study concentration-dependent dynamics since each molecule can be observed under controlled concentration conditions.
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 reduces costs and complexity while enabling the analysis of single molecules at natural concentrations, providing efficient sequencing and SNP detection with improved signal-to-noise ratio and throughput.
Implementation Method 1
a light source configured to illuminate the sample holder, an optical assembly configured to collect and detect separately and substantially simultaneously light emissions associated with the plurality of light emitting analytes
Implementation Method 2
an optical assembly configured to collect and detect separately and substantially simultaneously light emissions associated with the plurality of light emitting analytes
Data Source
AI summary
The present disclosure provides apparatus, systems and method for detecting separately and substantially simultaneously light emissions from a plurality of localized light-emitting analytes. A system according to exemplary embodiments of the present disclosure comprises a sample holder having structures formed thereon for spatially separating and constraining a plurality of light-emitting analytes each having a single nucleic acid molecule or a single nucleic acid polymerizing enzyme, a light source configured to illuminate the sample holder, an optical assembly configured to collect and detect separately and substantially simultaneously light emissions associated with the plurality of light emitting analytes. The system may further include a computer system configured to analyze the light emissions to determine the structures or properties of a target nucleic acid molecule associated with each analyte.