Split Nucleic Acid Labels for Specific High-Plex Proximity Detection
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Solution Overview
Problem
Existing biological sample analysis methods face challenges with cross-reactivity of affinity reagents, leading to false-positive results and reduced specificity, particularly in high-plex assays, and are limited by high background autofluorescence in tissue samples.
Innovation Solution
A label comprising a first and second nucleic acid backbone with labelling moieties and a guest molecule that forms a complex under specific conditions, allowing controlled proximity hybridization (hgPHA) to detect analyte proximity with high specificity and flexibility, compatible with enzymatic and non-enzymatic amplification.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If affinity reagents are used for detecting analytes in high-plex assays, then multiplexing capability is improved, but cross-reactivity increases leading to false-positive results
Solution Approach 1:
The label is divided into two separate nucleic acid backbones (first and second) that are spatially separated and can be independently controlled. This segmentation allows the system to distinguish between specific analyte binding events and non-specific cross-reactivity, thereby maintaining high specificity in high-plex assays while preserving multiplexing capability.
Solution Approach 2:
The patent introduces a controlled proximity hybridization mechanism as an intermediary step between analyte binding and signal generation. The two nucleic acid backbones act as intermediaries that only come into proximity when the affinity reagents are bound to adjacent analytes, providing an additional layer of verification that reduces false-positive results.
2Reliability
If conventional labels are used for detecting analytes, then detection capability is provided, but background autofluorescence in tissue samples increases
Solution Approach 1:
The patent employs fluorescent labels that undergo changes in their emission characteristics upon hybridization of the two nucleic acid backbones. This color change mechanism allows the signal to be differentiated from the static background autofluorescence of tissue samples, improving the signal-to-noise ratio and detection capability.
Solution Approach 2:
The label's optical parameters (such as fluorescence emission wavelength or intensity) are changed dynamically based on the hybridization state of the nucleic acid backbones. This parameter change enables the detection signal to be distinguished from the constant background autofluorescence, thereby reducing the harmful effect of background noise.
3Adaptability or versatility
If multiple markers are handled separately, then flexibility of use is maintained, but complexity of handling increases
Solution Approach 1:
The patent merges the control and detection functions into a single integrated label structure comprising two nucleic acid backbones and associated affinity reagents. This merging allows multiple markers to be handled as unified units that can be simultaneously controlled and detected, reducing the complexity of handling while maintaining flexibility through the modular nature of the nucleic acid components.
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
Enhances assay specificity by mitigating cross-reactivity and autofluorescence, enabling efficient detection of analyte proximities with rapid and easy readouts, suitable for high-plex molecular interaction analysis.
Implementation Method 1
The first nucleic acid backbone and the second nucleic acid backbone are configured to hybridise at least partially to each other
Implementation Method 2
at least one guest molecule configured to form a complex with a host molecule
Data Source
AI summary
A label for analyzing a biological sample includes a first label part comprising a first nucleic acid backbone, and a second label part comprising a second nucleic acid backbone. The first nucleic acid backbone and the second nucleic acid backbone are configured to hybridise at least partially to each other. The label further includes at least one first labelling moiety and at least one second labelling moiety, and at least one guest molecule configured to form a complex with a host molecule.