Fluorogenic Nucleosides for Low-Background Target Detection
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Solution Overview
Problem
Existing methods for labeling molecular targets such as small molecules, proteins, and nucleic acids with fluorescent probes face challenges due to high background fluorescence, leading to poor imaging agents and suboptimal results in applications like sequencing, biosensing, drug delivery, and microscopy.
Innovation Solution
Development of fluorogenic nucleosides, including fluorogenic nucleoside triphosphates, which undergo a change in fluorescence intensity and lifetime upon specific events like target binding, enabling the use of fluorogenic oligonucleotides as probes for target detection.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If fluorescent probes are used to label molecular targets, then labeling can be achieved, but high background fluorescence occurs making real-time observation difficult
Solution Approach 1:
The patent applies parameter changes by modifying the fluorescence properties of nucleoside triphosphates through chemical structure modification. The fluorogenic NTPs are designed to have suppressed fluorescence in the absence of target binding, and undergo a switch to high fluorescence emission upon binding to the target molecule. This changes the fluorescence parameter from constant high emission to conditional emission based on binding state.
Solution Approach 2:
The invention implements dynamics by creating a fluorescent probe whose emission properties are not static but dynamically change in response to target binding. The fluorogenic NTP transitions from a non-fluorescent or low-fluorescent state to a highly fluorescent state upon binding, enabling real-time detection of the binding event without requiring washing steps.
2Reliability
If conventional fluorescent tags are used in DNA and RNA oligonucleotides, then labeling is achieved, but imaging performance deteriorates due to high background fluorescence
Solution Approach 1:
The patent modifies the fluorescence parameter of oligonucleotide labels by incorporating fluorogenic NTPs that undergo a fluorescence switch upon target binding. This changes the illumination intensity from continuously high background fluorescence to selective high fluorescence only when target binding occurs, significantly improving imaging performance.
3Measurement precision
If fluorescent probes are used for target detection, then detection capability is provided, but washing steps are required increasing protocol complexity
Solution Approach 1:
The fluorogenic NTP probe performs self-service by automatically providing the signal change that indicates target binding. The probe itself undergoes the fluorescence switch in response to binding, eliminating the need for external washing steps or additional processing steps to separate bound from unbound probe. The binding event itself generates the detectable signal.
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 fluorogenic probes provide enhanced imaging capabilities by reducing background fluorescence and allowing real-time labeling without washing steps, improving the effectiveness of sequencing, biosensing, and microscopy applications.
Implementation Method 1
Fluorogenic probes are a class of chemical sensors that undergo a change (e.g., increase) in their fluorescence emission intensity and/or fluorescence lifetime upon the occurrence of a particular physical or chemical event
Data Source
AI summary
Provided herein are Anorogenic nucleosides (e.g., Anorogenic nucleoside triphosphates (NTPs), e.g., Anorogenic reversible terminator nucleoside triphosphates) which can be used in the synthesis of Anorogenic oligonucleotides (e.g., Anorogenic DNA or RNA oligonucleotides, such as Anorogenic RNA aptamers). The Anorogenic oligonucleotides (e.g., Anorogenic DNA or RNA oligonucleotides, such as, Anorogenic RNA aptamers) can be used as Anorogenic probes to detect targets (e.g., antigens, biomarkers).