Microfluidic Droplet System for Single-Cell Phenotype Genotype Coupling
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Solution Overview
Problem
Current microfluidic methods face challenges in accurately combining phenotypic and genotypic screenings at the single cell level, particularly in recovering specific cell genotypes and phenotypes, and in efficiently mixing reagents within microfluidic devices, which limits the ability to accurately assign phenotypes to genotypes.
Innovation Solution
A microfluidic system comprising a solid support with spatially separated groups of oligonucleotides, where cells are encapsulated in droplets and fused with reagent droplets containing lysis compositions, allowing oligonucleotides to attach to nucleic acids for phenotypic and genotypic analysis, enabling precise determination of phenotypes and genotypes.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If multiple reporter systems are multiplexed to increase the number of readouts for phenotypic analysis, then the number of simultaneous measurements is improved, but the costs and time for screening increase
Solution Approach 1:
The invention segments the phenotypic analysis into multiple independent droplets, each containing a single cell and specific reagents. This allows parallel processing of multiple phenotypic readouts simultaneously in separate droplets, increasing throughput without proportionally increasing the time required for each individual measurement.
Solution Approach 2:
The invention nests multiple analysis steps within a single droplet by sequentially adding reagents and performing operations. The droplet serves as a contained reaction chamber where multiple phenotypic assays can be performed in sequence, allowing the system to maintain high throughput while completing comprehensive analyses.
2Measurement precision
If phenotypic and genotypic analyses are performed at the single cell level, then the accuracy of phenotype-genotype assignment is improved, but the difficulty of recovering specific cell genotypes and phenotypes increases
Solution Approach 1:
The invention introduces barcoded magnetic beads as intermediaries that simultaneously carry both phenotypic information (via fluorescent reporters) and genotypic information (via barcodes). These beads mediate the linkage between phenotype and genotype by being internalized by cells and allowing subsequent retrieval and sequencing of barcode information, thus solving the difficulty of recovering specific cell genotypes while maintaining high accuracy in assignment.
3Device complexity
If reagents are mixed in bulk before compartmentalization, then the mixing process is simplified, but the initial reaction products fail to co-localize with their initiating target
Solution Approach 1:
The invention performs preliminary compartmentalization of reagents into separate droplets before initiating reactions. Each droplet contains specific reagents pre-grouped together, ensuring that when the assay is triggered, the reagents are already in the correct spatial arrangement to produce co-localized reaction products. This preliminary organization eliminates the need for complex post-compartmentalization mixing while ensuring reliable co-localization.
4Ease of operation
If post_analysis retrieval of individual samples is performed, then the ability to access specific droplets is improved, but the retrieval process becomes difficult to achieve
Solution Approach 1:
The invention uses barcoded magnetic beads as retrievable intermediaries. After the phenotypic and genotypic analyses are completed within the droplets, the magnetic beads can be selectively retrieved using magnetic fields based on their barcode sequences. This allows easy access to individual samples through magnetic separation while avoiding complex mechanical retrieval mechanisms, thus improving ease of operation without significantly increasing device complexity.
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 precise coupling of phenotype and genotype information at the single cell level, improving the accuracy of phenotypic and genotypic screenings and enabling reliable assignment of phenotypes to genotypes, while also facilitating high-throughput analysis and efficient reagent mixing.
Implementation Method 1
droplets of a second type, comprising reagents for performing one or more reactions are injected into and/or flowed through a channel of the microfluidics device, such that the droplets of a second type may get trapped separately inside each of the reservoirs of the microfluidics device. Consequently, each reservoir of the microfluidics device comprises one droplet of a first type and one droplet of a second type. A droplet of a first type may be fused with a droplet of the second type
Data Source
AI summary
The invention relates to a microfluidic system comprising: a) a solid support comprising at least a first group of oligonucleotides, i. wherein each oligonucleotide in said group comprises a nucleic acid sequence of a first type, of a second type and/or a further type, ii. wherein said nucleic acid sequence of a first type is a barcode sequence, iii. and oligonucleotides comprising the same barcode sequence are grouped together in a group of oligonucleotides on said solid support, iv. wherein the first and further oligonucleotide groups are spatially separated on said solid support, b) wherein said one or more groups of oligonucleotide groups on said solid support are within separate reservoirs of the microfluidics system, c) wherein the one or more reservoirs are accessible to fluids, cells, chemicals and/or microdroplet by means of channels, and d) wherein each reservoir comprises comprising a group of oligonucleotides on said solid support is also trap for a microfluidic droplet.