Automated eDNA Quantification Apparatus with Capture Probes
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Solution Overview
Problem
Current methods for analyzing environmental DNA (eDNA) face challenges in accurately detecting and quantifying cell-free eDNA due to its rapid decay and stabilization issues, often requiring complex sample preparation and handling, which can lead to loss and interference from background molecules.
Innovation Solution
A fieldable apparatus and method for automatically collecting and quantifying eDNA in environmental samples using a system that includes a sample inlet, reagent storage, and a digital droplet PCR instrument, which measures eDNA levels without dissociating target DNA from cells or background molecules, enabling accurate detection of cell-free eDNA before decay without stabilization methods.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If complex sample preparation and handling procedures are used to detect and quantify cell-free eDNA, then detection capability is improved, but sample loss and interference from background molecules increase
Solution Approach 1:
The system performs preliminary actions by pre-coating the collection filter with capture probes before sample introduction, and by pre-assembling the complete analysis workflow in a automated manner. This ensures that when cell-free eDNA molecules pass through the filter, they are immediately captured and processed, minimizing sample loss and background interference throughout the analysis process.
2Measurement precision
If complex sample preparation and handling procedures are used to detect and quantify cell-free eDNA, then detection capability is improved, but interference from background molecules increases
Solution Approach 1:
The system extracts only the target cell-free eDNA molecules from the complex environmental sample matrix by using capture probes that specifically bind to target sequences. The filter architecture allows non-target background molecules to pass through while retaining only the captured eDNA, effectively separating the signal of interest from interfering background components.
Solution Approach 2:
The capture probes coated on the filter serve as intermediaries that specifically recognize and bind to target eDNA sequences. These probes mediate the selective capture process, enabling the system to distinguish target molecules from background noise without requiring complex sample preparation steps that would otherwise be needed to reduce interference.
3Ease of operation
If cell-free eDNA is stabilized using reagents, then sample handling is improved, but quantification accuracy deteriorates due to loss and interference
Solution Approach 1:
The system enables self-service by performing automated sample collection, filtration, capture, and analysis without requiring external stabilization reagents or manual intervention. The integrated workflow automatically processes samples from collection through quantification, eliminating the need for separate stabilization steps that would otherwise be required to maintain sample integrity during handling.
4Productivity
If manual collection and processing steps are used, then sample analysis is achieved, but processing time and potential for loss increase
Solution Approach 1:
The system merges multiple discrete manual operations into a single integrated automated instrument that performs sample collection, filtration, molecular capture, and quantitative analysis in one continuous workflow. This consolidation eliminates transfer steps between different equipment and operations, thereby reducing both processing time and the risk of sample loss during manual handling.
Solution Approach 2:
The apparatus is designed as a universal multi-functional system that can handle various sample types and perform complete eDNA analysis workflows within a single device. This multi-functionality eliminates the need for multiple specialized instruments and manual transfer operations, significantly reducing processing time while maintaining comprehensive sample analysis capability.
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 apparatus allows for continuous, accurate measurement of eDNA levels in environmental samples with reduced sample handling, minimizing losses and interference, and enables precise quantification of gene copy numbers without the need for extensive sample preparation or stabilization, enhancing detection accuracy and simplifying the collection process.
Implementation Method 1
digital droplet PCR instrument, which measures eDNA levels
Implementation Method 2
measures eDNA levels without dissociating target DNA from cells or background molecules
Data Source
AI summary
A fieldable processing and detection apparatus for automatically collecting, preparing, Identifying and quantifying environmental DNA in samples of a material of Interest. Environmental samples of materials of interest are combined with polymerase chain reaction (PGR) reagents that are selectively compatible with the material of Interest and mixed. Various processing methods may be employed at the discretion of the operator and include droplet concentration, thermoprofiling, particle separation and other techniques or methods that are compatible with and suitable for the specific material of interest and the testing environment The system will selectively lyse cells, breaking down the cell membrane via mechanical disruption, ultrasound, thermocycling, or other suitable techniques and thereafter quantify the preamplified concentration of target nucleic add sequences using digital quantification.