Electrochemical Biosensor Single-Step Detection
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Solution Overview
Problem
Current electrochemical protein biosensors are challenging to operate in a sample-in-answer-out (SIAO) manner, especially with unprocessed clinical samples, due to their dependence on multiple steps involving washes and reagent additions, which complicates their use in point-of-care protein analysis.
Innovation Solution
The electrochemical bio-barcode assay integrates biorecognition with signal transduction using molecular machines and nanostructured electrodes, allowing for single-step analysis by releasing a detectable label close to the electrode surface upon target analyte binding, eliminating the need for multi-step processing.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If multi-step processing with washes and reagent additions is used, then detection sensitivity is improved, but device complexity and ease of operation deteriorate
Solution Approach 1:
The patent combines multiple processing steps into a single integrated reaction well. The sample, capture probes, and detection probes are all present in the same well simultaneously, allowing target capture and signal generation to occur together without sequential washes or reagent additions. This merging eliminates operational complexity while preserving detection sensitivity through the integrated assay design.
Solution Approach 2:
The detection probe serves multiple functions: it captures the target analyte through its recognition element, generates the signal through its reporter moiety, and undergoes toehold-mediated strand displacement for signal amplification. This multi-functionality eliminates the need for separate capture and detection steps, reducing operational complexity while maintaining high detection sensitivity.
2Measurement precision
If multi-step processing with washes and reagent additions is used, then detection sensitivity is improved, but ease of operation deteriorates
Solution Approach 1:
The patent combines multiple processing steps into a single integrated reaction well. The sample, capture probes, and detection probes are all present in the same well simultaneously, allowing target capture and signal generation to occur together without sequential washes or reagent additions. This merging eliminates operational complexity while preserving detection sensitivity through the integrated assay design.
Solution Approach 2:
The assay system performs self-service through toehold-mediated strand displacement, where the detection probe automatically undergoes conformational change and releases the reporter moiety upon target binding, without requiring external intervention or multiple processing steps. This self-service mechanism simplifies operation while maintaining detection sensitivity.
3Measurement precision
If loaded nanoparticles or enzymes are used for amplifying the nucleic acid reporter, then detection sensitivity is improved, but device complexity deteriorates
Solution Approach 1:
The patent uses a toehold sequence as an intermediary element that mediates the strand displacement reaction. The toehold allows the detection probe to bind to the captured target and subsequently undergo conformational change, releasing the reporter moiety without requiring nanoparticles or enzymes. This intermediary mechanism provides signal amplification while avoiding the operational complexity of additional reagents.
Solution Approach 2:
The patent replaces the mechanical/enzymatic amplification system (nanoparticles or enzymes) with a purely nucleic acid-based toehold-mediated strand displacement mechanism. This substitution eliminates the need for additional amplification reagents and steps while maintaining detection sensitivity through the inherent amplification capability of the strand displacement reaction.
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 enables sensitive and specific protein detection in undiluted human plasma with a log-linear range of 1 ng/mL-200 ng/mL and a limit of detection (LOD) of 0.4 ng/mL, suitable for clinical decision-making at the point-of-care, while reducing operational complexity.
Implementation Method 1
the recognition moiety of the first detection probe binds to the target analyte, and the recognition moiety of the second detection probe binds to a different portion of the target analyte
Implementation Method 2
the first portion of the first detection probe binds by complementarity to the overhang of the first strand of the double-stranded oligonucleotide, and the first portion of the second detection probe binds by complementarity to an internal segment of the first strand
Implementation Method 3
signal readout using, for example, nanostructured, electrodes
Data Source
AI summary
This disclosure relates to a biosensor for detecting a target analyte in a sample comprising: a double-stranded oligonucleotide comprising an overhang on a first strand of the oligonucleotide, and a second strand of the oligonucleotide that is a reporter moiety comprising a detectable label; a first detection probe comprising a recognition moiety and a junction forming moiety, wherein the junction forming moiety comprises a first portion capable of binding by complementarity to the overhang of the first strand of the double-stranded oligonucleotide; a second detection probe comprising a recognition moiety and a junction forming moiety, wherein the junction forming moiety comprises a first portion capable of binding by complementarity to an internal segment of the first strand of the double-stranded oligonucleotide; and a capture probe functionalized on an electrode, wherein the capture probe comprises an immobilized strand attached to the electrode. Methods and uses thereof are also disclosed herein.