Dynamic DNA Complexes for Tunable Signal Amplification
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Solution Overview
Problem
Current molecular imaging techniques face challenges in controlling signal amplification levels and balancing the intensities of multiple fluorophores, leading to limited control over reporting levels and increased noise from spectral overlap, which complicates the detection of dilute molecular targets in biological samples.
Innovation Solution
The use of dynamic DNA complexes that can generate structurally-organized, branched or dendritic reporting complexes with controlled dye coupling, allowing for tunable reporting intensities and color-time-sequence coding to identify and localize multiple molecular targets without chemical or optical treatments, enabling precise labeling and imaging of proteins and biomacromolecules.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If enzymatic signal amplification strategies (TSA, RCA) are used to detect low level molecular targets, then detection sensitivity is improved, but control over final amplification levels is limited
Solution Approach 1:
The patent applies parameter changes by using non-enzymatic chemical reactions with tunable kinetics to replace enzymatic amplification. The chemical environment and reaction conditions can be precisely controlled to regulate amplification levels, achieving both high detection sensitivity and controllable signal intensity for multiplexed imaging.
Solution Approach 2:
The invention implements local quality by creating spatially and temporally controlled amplification zones. Different regions of the sample can have different amplification levels achieved through localized application of reagents or controlled reaction timing, allowing precise regulation of signal intensity in specific areas while maintaining high sensitivity.
2Adaptability or versatility
If multiple fluorophores are used for multiplexed molecular imaging, then the ability to detect multiple molecular targets is improved, but spectral overlap increases leading to noise and signal bleed-through
Solution Approach 1:
The patent employs periodic action through sequential or alternating imaging cycles. Different fluorophore combinations are imaged at different time points, allowing spectral unmixing and separation of overlapping signals. This temporal multiplexing approach enables detection of multiple targets while avoiding spectral interference.
Solution Approach 2:
The invention adds the time dimension to multiplexed imaging by using color-time-sequence codes. Targets are not only distinguished by spectral color but also by the temporal sequence of their appearance or intensity changes, creating a multi-dimensional detection space that separates overlapping signals both spectrally and temporally.
3Quantity of substance
If chemical reactions are used for signal amplification, then amplification gain is achieved, but reaction rates depend on local concentrations and chemical environment leading to sample-dependent variability
Solution Approach 1:
The patent introduces an intermediary chemical environment that mediates the amplification reaction. By controlling the buffer conditions, pH, and chemical composition of the reaction medium, the amplification gain becomes decoupled from variations in local sample concentrations, ensuring uniform and reliable signal amplification across different samples and 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 allows for highly-multiplexed molecular analyses with controlled reporting intensities and precise localization of multiple targets, reducing noise and enhancing detection sensitivity, enabling visualization of numerous targets within a single sample without compromising spatial resolution.
Implementation Method 1
The dynamic DNA complexes are reacted via strand displacement with branched or dendritic dynamic DNA complexes to create dynamic DNA-target complexes
Implementation Method 2
The probe comprises at least one fluorescent dye and a polynucleotide
Data Source
AI summary
The present invention generally relates to methods for detecting a target in a sample; methods for modulating the reporting intensity of a labeled target in a sample of fixed cells or tissues; methods for detecting the location of at least two targets in a sample; and related compositions.