Probe Chain Structure for Specific Molecular Detection
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Solution Overview
Problem
Current molecular biological methods, such as FISH, are limited in detecting target sequences with low copy numbers and shorter half-lives, like mRNA, and struggle to differentiate between living and dead microorganisms, especially at the species or pathogenicity-specific levels, due to variability in rRNA content and signal strength.
Innovation Solution
A probe complex is designed with primary and secondary probes that form a chain structure, allowing for signal amplification through complementary binding, enabling the detection of low-copy nucleic acid or amino acid sequences, including mRNA and species-specific DNA, using optically detectable labels for enhanced sensitivity and specificity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional FISH methods use rRNA as target substance, then high signal strength is achieved, but differentiation between species and live/dead status becomes unreliable due to varying rRNA content
Solution Approach 1:
The detection system is segmented into multiple independent probe types (primary probes targeting different sequences), each capable of detecting specific target substances. This segmentation allows simultaneous detection of multiple markers (rRNA, mRNA, DNA) to reliably differentiate between live/dead status and identify species-specific sequences, resolving the contradiction between detection specificity and reliability
Solution Approach 2:
The invention changes the detection parameters by targeting multiple different nucleic acid sequences (rRNA, mRNA, DNA) with different half-lives and copy numbers. By using probes with different target specificities and combining their signals, the system achieves both high detection precision and reliable live/dead differentiation
2Reliability
If FISH methods target sequences with short half-life and low copy numbers, then better live/dead differentiation is achieved, but signal strength becomes insufficient for reliable detection
Solution Approach 1:
The invention merges signals from multiple probe types targeting different sequences (rRNA, mRNA, DNA) to achieve sufficient overall signal strength. By combining detection results from multiple markers with different copy numbers and half-lives, the system maintains high detection sensitivity while enabling reliable live/dead differentiation through pattern recognition
Solution Approach 2:
The method performs preliminary detection of multiple target sequences simultaneously using specifically designed probes. By pre-coating beads with multiple probe types and detecting their combined signals, the system achieves both high sensitivity for low-copy sequences and reliable differentiation accuracy
3Reliability
If enzyme-based methods are used for live/dead differentiation, then living stages can be detected, but dead microorganisms may still show positive signals complicating analysis
Solution Approach 1:
The invention introduces nucleic acid probes as intermediary markers that directly bind to specific sequences (mRNA, DNA) rather than relying on enzyme activity. These nucleic acid targets persist differently in live versus dead cells, providing a more specific mediator for differentiation that avoids the false positive problem of enzyme-based methods
4Adaptability or versatility
If multiple target sequences are detected in one procedure, then comprehensive analysis is achieved, but cost and procedural complexity increase
Solution Approach 1:
The invention creates a universal detection platform where a single assay can detect multiple target sequences (rRNA, mRNA, DNA) using a standardized protocol. The same basic FISH procedure and detection methodology works for all target types, achieving multi-functionality without proportionally increasing procedural complexity or cost
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 approach allows for the specific detection of low-copy target sequences, improving differentiation between living and dead microorganisms and enabling the detection of species- or pathogenicity-specific sequences, with increased fluorescence intensity and signal-to-noise ratio, suitable for use outside laboratory environments.
Implementation Method 1
the probe complex binds specifically to a target segment of the substance... the probe complex has at least one probe type as a primary probe with a target-specific segment that is complementary to the target segment of the substance
Implementation Method 2
an interaction with at least one detectable marker can trigger a preferably optically detectable signal
Data Source
Figure 1
AI summary
It is proposed to provide a probe complex (1) for the specific detection of at least one substance (2), wherein the probe complex (1) binds specifically to a target segment (7) of the substance (2) and wherein an interaction with at least one detectable marker (8) can trigger a preferably optically detectable signal, wherein the probe complex (1) comprises at least one probe type as a primary probe (6) with a target-specific segment (5) and at least one further probe type as a secondary probe (9) with a target-specific segment (10), wherein the target-specific segments of all probe types of the probe complex (1) are coordinated such that a chain (3) of at least three probes (4) can be formed.