Covalent Fluorescent Probe for Stable In Vivo Target Detection

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Solution Overview

Problem

Current fluorescent probes for detecting target molecules are limited by the separation of their components during detection, leading to biological contamination and high detection thresholds, especially in vivo applications, and require additional handling steps like injection and washing.

Innovation Solution

A fluorescent probe with a receptor bonded to a polypeptide via a covalent bond, forming a FRET donor/acceptor pair with fluorochromes, which remains intact during detection, eliminating separation risks and reducing handling operations.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Reliability

If a fluorescent probe uses non-covalent bonding between antibody and protein A, then the probe can be assembled, but the components separate during detection causing biological contamination and high detection thresholds

Engineering Contradiction:

Improvestability of probe componentsVSAvoidbiological contamination

Core Design Contradiction:

ReliabilityVSObject-affected harmful factors

Solution Approach 1:

The patent applies preliminary action by establishing covalent bonds between the antibody and protein A during probe assembly, before the detection process begins. This pre-established strong chemical bonding prevents component separation during subsequent detection operations, eliminating the reliability issue and preventing biological contamination that would otherwise occur.

Inventive Principle:

Principle #10Preliminary action

Solution Approach 2:

The patent changes the bonding parameter from non-covalent (weak, reversible) to covalent (strong, irreversible) between the antibody and protein A. This parameter change fundamentally alters the stability of the probe components, ensuring they remain bound during detection and do not separate to cause contamination or increase detection thresholds.

Inventive Principle:

Principle #35Parameter changes

2Measurement precision

If a fluorescent probe uses two separate antibodies labelled with fluorochromes, then the probe can be constructed, but intramolecular FRET effect cannot be achieved reducing detection sensitivity

Engineering Contradiction:

Improvedetection sensitivityVSAvoidprobe structure

Core Design Contradiction:

Measurement precisionVSDevice complexity

Solution Approach 1:

The patent merges two separate antibody components into a single integrated fluorescent probe structure by covalently binding them through protein A. This combining allows the fluorochromes attached to each antibody to be positioned in close proximity, enabling the intramolecular FRET effect that significantly improves detection sensitivity compared to separate antibody approaches.

Inventive Principle:

Principle #5Merging (Combining)

Solution Approach 2:

The patent uses protein A as an intermediary component that covalently links the two antibodies together. This mediator enables the formation of a unified probe structure where the fluorochromes can interact through FRET, while protein A itself remains part of the stable covalent structure, facilitating both the structural integration and the optical interaction.

Inventive Principle:

Principle #24Intermediary (Mediator)

3Ease of operation

If fluorescent probes are injected in vivo, then detection can be performed, but safety must be demonstrated and biological contamination risk increases

Engineering Contradiction:

Improvedetection accessibilityVSAvoidsafety and contamination risk

Core Design Contradiction:

Ease of operationVSObject-affected harmful factors

Solution Approach 1:

The patent applies preliminary anti-action by using covalent bonding to prevent the separation of probe components before in vivo injection. By ensuring the antibody and protein A remain firmly bound through strong chemical bonds, the probe maintains its structural integrity during injection and in vivo circulation, preventing the release of free antibodies or components that would cause biological contamination or safety issues.

Inventive Principle:

Principle #9Preliminary anti-action

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 design enhances sensitivity and stability, allowing for rapid and accurate detection of target molecules without biological contamination, suitable for both in vitro and in vivo use, including during surgery or endoscopy, with reduced handling and improved safety.

Implementation Method 1

The detection of target molecules using FRET technology, based on non-radiative energy transfer between two fluorochromes

Methodology Applied

Scientific EffectFRET (Förster resonance energy transfer):

Implementation Method 2

two fluorochromes Fa and Fb; wherein the fluorochrome Fa is bonded to the receptor and the fluorochrome Fb is bonded to the polypeptide; and the fluorochromes Fa and Fb form a FRET donor/acceptor pair

Methodology Applied

Scientific EffectFluorescence: Fluorescence

Data Source

PatentUS20230349910A1Fluorescent reporter and use thereof for the detection of target molecules

Publication Date: 2023.11.02 KAMAX INNOVATIVE SYST

AI summary

A device for detecting a target molecule and/or measuring the concentration of a target molecule, which includes: a substrate at the surface of which is covalently attached a grafting molecule; at least one fluorescent probe including at least one receptor bonded to a polypeptide via a covalent bond; two fluorochromes Fa and Fb, in which the fluorochrome Fa is bonded to the receptor and fluorochrome Fb is bonded to the polypeptide; and the fluorochromes Fa and Fb form a FRET donor/acceptor pair, in which the polypeptide is bonded to the grafting molecule via a covalent bond. Also, a fluorescent probe and a method for detecting a target molecule and/or measuring the concentration of a target molecule.