Nucleic Acid Detection Apparatus Using Segmented Compartments
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Solution Overview
Problem
Current methods for detecting low-concentration nucleic acids using CRISPR-Cas technology are complex and time-consuming, often requiring amplification steps that complicate the detection process.
Innovation Solution
A nucleic acid detection apparatus and method that distributes a sample and detection reagent to individual independent separated compartments, utilizing an effector protein, crRNA, and a reporter molecule to generate fluorescence, allowing for direct detection of nucleic acids without amplification, using a distribution unit, activation unit, fluorescence generation unit, and identification unit to determine fluorescence intensity and identify compartments with detectable signals.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If amplification steps are performed to detect low-concentration nucleic acids, then detection sensitivity is improved, but device complexity and detection time increase
Solution Approach 1:
The system divides the sample into multiple individual independent separated compartments (microdroplets or microwells), each containing the detection reagents. This segmentation allows direct detection of low-concentration nucleic acids without amplification by concentrating target molecules in discrete compartments, resolving the contradiction between detection sensitivity and operational simplicity.
Solution Approach 2:
The invention changes the physical state and concentration parameters by distributing the sample into numerous small compartments, effectively concentrating target nucleic acids in each compartment without requiring amplification. This parameter change enables direct detection while maintaining simplicity.
2Measurement precision
If amplification steps are performed to detect low-concentration nucleic acids, then detection sensitivity is improved, but detection time increases
Solution Approach 1:
By segmenting the sample into many independent compartments, the system enables parallel processing of multiple sample portions simultaneously. This allows direct detection of low-concentration targets without time-consuming amplification steps, reducing total detection time while maintaining sensitivity.
Solution Approach 2:
The system uses an excessive number of compartments relative to the amount of target nucleic acid, ensuring that even low-concentration samples will have sufficient target molecules in some compartments to generate detectable signals, eliminating the need for amplification and reducing detection time.
3Ease of operation
If individual independent separated compartments are used for direct detection, then operation simplicity is improved, but detection sensitivity may decrease
Solution Approach 1:
The segmentation into numerous independent compartments actually improves detection sensitivity through statistical effects - with enough compartments, the probability of finding target molecules increases, enabling direct detection without amplification while maintaining simplicity.
Solution Approach 2:
By changing the compartment size and number parameters, the system achieves optimal conditions for direct detection, where the concentration of target molecules in each small compartment is sufficient for detection without amplification, maintaining both simplicity and sensitivity.
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 simple and rapid detection of low-concentration nucleic acids by generating fluorescence in compartments where the target nucleic acid is present, eliminating the need for amplification steps and reducing detection time, while maintaining high sensitivity.
Implementation Method 1
Cas12a cleaves the single-stranded DNA of the reporter molecule by a trans-cleavage reaction. Thus, the fluorescent substance and the quencher are separated, and fluorescence is generated.
Implementation Method 2
when a reporter molecule in which a fluorescent substance and a quencher are linked to each other by single-stranded DNA is added to a reaction system, Cas12a cleaves the single-stranded DNA of the reporter molecule by a trans-cleavage reaction. Thus, the fluorescent substance and the quencher are separated, and fluorescence is generated.
Data Source
AI summary
Provided is a nucleic acid detection apparatus including: a distribution unit configured to distribute a sample and a detection reagent to a plurality of individual independent separated compartments, the sample containing a target nucleic acid, and the detection reagent containing an effector protein, crRNA to be bound to the target nucleic acid, and a reporter molecule; an activation unit configured to activate the effector protein through binding of the crRNA to the target nucleic acid; a fluorescence generation unit configured to modify the reporter molecule with the activated effector protein to generate fluorescence; a fluorescence detection unit configured to detect the fluorescence; and an identification unit configured to determine, based on a detection result obtained with the fluorescence detection unit, a fluorescence intensity of each of the individual independent separated compartments, and to identify each of the individual independent separated compartments having a fluorescence intensity exceeding a predetermined threshold value.