Multi-dye Chromogenic Conjugates for IHC Staining
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Solution Overview
Problem
Existing methods for immunohistochemistry (IHC) and in situ hybridization (ISH) face challenges in robustly detecting low-abundance cellular markers while minimizing non-specific background signals, as current signal amplification techniques like Tyramide Signal Amplification (TSA) and Quinone Methide Signal Amplification (QMSA) can lead to increased background noise, obscuring faint but clinically significant signals.
Innovation Solution
Development of multi-dye conjugates comprising a tissue-reactive moiety and at least two chromophores, such as quinone methide precursors or tyramides, which are conjugated through multi-functional linkers, allowing for distinct and enhanced signal generation without increasing background noise.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If signal amplification techniques like TSA or QMSA are used to detect low-abundance cellular markers, then detection sensitivity is improved, but background noise increases obscuring faint signals
Solution Approach 1:
The patent combines multiple dye molecules (e.g., tyramide and quinone methide) into a single conjugate molecule, allowing simultaneous deposition at the target site through a single enzymatic reaction. This merging approach ensures that both dyes are deposited in equal amounts and locations, providing robust signal amplification for low-abundance markers while avoiding the background noise problems associated with separate dye applications.
Solution Approach 2:
The invention creates composite conjugate molecules that integrate multiple chromogenic components (tyramide and quinone methide) with different detection properties into a single molecular entity. This composite structure enables coordinated deposition of multiple signals at the target site, enhancing detection sensitivity while maintaining signal specificity and reducing background interference through synchronized deposition.
2Adaptability or versatility
If multiple separate dye conjugates are used to achieve multi-color detection, then detection versatility is improved, but deposition uniformity deteriorates due to varying reaction rates
Solution Approach 1:
The patent merges multiple dye molecules into a single conjugate structure where different dyes (e.g., tyramide and quinone methide) are covalently linked together. This ensures that when the conjugate undergoes enzymatic deposition, all dye molecules are deposited simultaneously and uniformly at the target site, eliminating the deposition uniformity issues that arise from using separate dye conjugates with different reaction kinetics.
Solution Approach 2:
The invention implements local quality by positioning different dye molecules at specific locations within the conjugate structure, allowing each dye to contribute its unique detection properties while ensuring uniform co-deposition. The spatial arrangement of dyes within the conjugate enables multi-color detection capability while maintaining consistent deposition patterns across all dye types.
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
The multi-dye conjugates provide higher overall signal intensity and distinct color outputs, overcoming the limitations of single-dye conjugates by ensuring even deposition and eliminating variations in color due to varying reaction rates, thus enhancing the detection of low-abundance targets in biological samples.
Implementation Method 1
contacting the biological sample with a labeling conjugate that comprises an enzyme; the enzyme converts the multi-dye conjugate to a reactive intermediate that covalently binds to the biological sample proximally to or directly on the target
Implementation Method 2
the short lifetime of the radical intermediate results in covalent binding of the tyramide to proteins on tissue in close proximity to the site of generation, giving discrete and specific signal
Data Source
AI summary
Disclosed herein are novel chromogenic conjugates, the conjugates comprising at least two detectable moieties.