Integrated ISFET Chip for Amplification and Sequencing
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Solution Overview
Problem
Current nucleic acid sequencing methods require spatial separation of amplification and sequencing steps, leading to increased costs, sample loss, and decreased sensitivity due to the need for skilled labor and separate equipment.
Innovation Solution
An integrated ion-sensitive apparatus and method that combines amplification and sequencing on a semiconductor chip with ISFETs, allowing for simultaneous clonal amplification and sequencing in the same well, using a microfluidic structure with heating means and removable seals to optimize conditions and prevent cross-contamination.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If spatial separation of amplification and sequencing steps is used, then each step can be optimized independently, but costs increase, sample loss increases, and sensitivity decreases
Solution Approach 1:
The patent merges the amplification and sequencing steps into a single integrated microfluidic device, allowing both processes to occur in the same well without spatial separation. This integration eliminates the need for separate amplification and sequencing machines, reducing system complexity while maintaining or improving sequencing sensitivity by preventing sample loss during transfer between devices.
2Productivity
If integrated amplification and sequencing is implemented, then hands-on time is reduced and sample loss is minimized, but device complexity increases
Solution Approach 1:
The integrated device is segmented into distinct functional zones within the microfluidic chip: an amplification zone with heating means for performing PCR or other amplification reactions, and a sequencing zone with ISFET sensors for detecting nucleotide incorporation. This segmentation allows each zone to be optimized for its specific function while being part of an integrated system, enabling high productivity without excessive device complexity.
Solution Approach 2:
The microfluidic device performs multiple functions within a single platform: it can perform nucleic acid amplification, hold amplified products, and conduct sequencing reactions without requiring transfer between devices. The ISFET sensors serve dual purposes by detecting both amplification progress and sequencing signals, demonstrating multi-functionality that increases productivity while managing device complexity.
3Measurement precision
If conventional fluorescent-based detection is used, then sequencing accuracy is maintained, but costs increase and safety concerns arise
Solution Approach 1:
The patent replaces the optical detection system (fluorescent microscopy and imaging equipment) with an electrical detection system using ISFET sensors. The ISFETs detect changes in ion concentration caused by nucleotide incorporation, converting a chemical/biological process into an electrical signal. This substitution eliminates the need for expensive optical instruments while maintaining detection accuracy and improving safety by removing the requirement for fluorescent labels and radioactive isotopes.
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 reduces hands-on time, minimizes sample loss, and increases the speed of the sequencing process, making it more suitable for commercial use by non-specialist professionals while maintaining high accuracy and sensitivity.
Implementation Method 1
an ISFET, which measures ion concentrations in solution, has been employed to detect nucleotide incorporation into a nucleic acid strand by detecting the change in hydrogen ion (H+, proton) concentration resulting from the reaction
Implementation Method 2
amplification means for amplifying the template polynucleotide on a surface of said chip and comprising at least one heating means suitable for conducting amplification of the template polynucleotide at temperatures elevated with respect to room temperature
Data Source
AI summary
Provided is a sensing apparatus comprising a chip for integrated amplification and sequencing of a template polynucleotide in a sample. The apparatus comprises a chip with at least one ISFET in a well or chamber, amplification means for amplifying the template polynucleotide on a surface of said chip and comprising at least one heating means suitable for conducting amplification of the template polynucleotide at temperatures elevated with respect to room temperature, and sequencing means for sequencing the amplified template polynucleotide in said well or chamber. Methods of use are also provided.