Oligonucleotide Probes for Rapid Bacterial Detection
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Solution Overview
Problem
Current methods for detecting bacterial pathogens are time-consuming, expensive, and often involve toxic or radioactive tracers, and lack sensitivity, especially for imaging bacterial colonization in humans.
Innovation Solution
Development of a probe comprising a substrate oligonucleotide with a fluorescence-reporter group and a fluorescence-quencher group separated by an RNAse- or DNAse-cleavable residue, which is cleaved by microbial nucleases, allowing for the detection of microbial infections through increased fluorescence.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If conventional methods are used for detecting bacterial pathogens, then detection can be performed, but the process is time-consuming and expensive
Solution Approach 1:
The patent replaces complex mechanical and chemical detection systems (culture methods, PCR, mass spectrometry) with a simple fluorescence-based optical detection system. The probe uses fluorophore-quencher pairs that generate fluorescent signals upon nuclease cleavage, enabling rapid detection without time-consuming culture steps or expensive equipment.
Solution Approach 2:
The patent changes the detection parameter from measuring bacterial growth (time-consuming) or genetic material amplification (expensive) to measuring fluorescence intensity (rapid and simple). The fluorescent signal provides direct, real-time information about bacterial presence and nuclease activity.
2Object-affected harmful factors
If conventional detection methods are used, then bacterial pathogens can be detected, but toxic or radioactive tracers are required
Solution Approach 1:
The patent uses non-toxic, biodegradable oligonucleotide probes with fluorophore-quencher pairs instead of persistent radioactive or toxic tracers. These probes are consumed in the detection process and can be safely eliminated from the body, eliminating long-term toxicity concerns while maintaining high detection reliability.
3Measurement precision
If conventional methods are used for imaging bacterial colonization, then detection is possible, but sensitivity is insufficient
Solution Approach 1:
The patent uses fluorescence emission (color change from non-fluorescent to fluorescent) as the detection signal. The fluorophore-quencher system provides high contrast signals that are easily detected by standard fluorescence imaging equipment, significantly improving sensitivity for imaging bacterial colonization compared to conventional methods.
4Illumination intensity
If FRET-based quenching is used, then fluorescence quenching occurs, but the emission spectrum of the fluorophore must overlap with the absorbance spectrum of the quencher
Solution Approach 1:
The patent employs universal fluorophore-quencher pairs (e.g., FAM-BHQ1, Cy5-BHQ3) that can be applied to multiple detection scenarios. The quencher molecules are designed to work with specific fluorophores across different spectral ranges, providing a standardized approach that simplifies probe design while maintaining high quenching efficiency.
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
Enables rapid, inexpensive, and non-toxic detection of microbial infections by measuring fluorescence changes in biological samples, providing a sensitive method for bacterial detection and imaging.
Implementation Method 1
a fluorescence-reporter group operably linked to the oligonucleotide
Implementation Method 2
FRET is one of the most common mechanisms of fluorescent quenching and can occur when the emission spectrum of the fluorescent donor overlaps the absorbance spectrum of the quencher and when the donor and quencher are within a sufficient distance known as the Forster distance
Data Source
AI summary
The present invention relates to a rapid detection of microbial-associated nuclease activity with chemically modified nuclease (e.g., ribonuclease) substrates, and probes and compositions useful in detection assays.