Microfluidic Microwell Array Integrating Cell Morphology and Gene Expression
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Solution Overview
Problem
Current methods lack a high-throughput approach to integrate gene expression profiles with morphological phenotypes at the single-cell level, often relying on bulk RNA-seq analysis or manual cell collection, which ignores cellular heterogeneity and has limited throughput.
Innovation Solution
A microfluidic device with a microwell array and interdigital electrode is used to trap individual cells, where capture oligonucleotides with unique barcode and identifier sequences capture mRNA, allowing for high-throughput integration of morphological characteristics and gene expression profiles through imaging and sequencing.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If bulk RNA-seq analysis is used, then analysis throughput is improved, but cellular heterogeneity is lost and measurement precision deteriorates
Solution Approach 1:
The invention segments the bulk sample into individual single cells using microfluidic droplet generation, where each droplet contains at most one cell. This segmentation enables simultaneous high-throughput processing of thousands of individual cells while maintaining cellular heterogeneity, as each cell's RNA is analyzed separately rather than being averaged in bulk.
Solution Approach 2:
The invention introduces barcoded beads as an intermediary element that captures and labels RNA from individual cells. Each bead carries a unique barcode that identifies the parent cell, allowing RNA from thousands of cells to be pooled and processed together while still enabling traceability to individual cells, thus achieving both high throughput and cellular resolution.
2Measurement precision
If manual cell collection using pipette is used, then measurement precision is improved, but productivity deteriorates
Solution Approach 1:
The invention replaces the manual mechanical pipetting process with an automated microfluidic system that uses flow-controlled droplet generation. This substitution maintains single-cell isolation precision while enabling automated high-throughput processing of thousands of cells, as the microfluidic system can continuously generate and process droplets without manual intervention.
Solution Approach 2:
The microfluidic system incorporates on-chip cell lysis and RNA capture functions, where the system automatically performs lysis buffer injection, RNA release, and bead-mediated RNA capture within each droplet without external manual operations. This self-service capability maintains single-cell resolution while dramatically increasing throughput by eliminating manual pipetting steps.
3Productivity
If high-throughput single-cell processing is implemented, then productivity is improved, but device complexity increases
Solution Approach 1:
The microfluidic chip is designed as a universal platform that integrates multiple functions including cell injection, droplet generation, on-chip cell lysis, RNA capture, and sample pooling in a single device. This multi-functionality achieves high-throughput single-cell processing while managing device complexity by consolidating operations that would otherwise require separate instruments and manual steps.
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 method achieves high single-cell purity, recovery rate, and sensitivity, with low RNA contamination, enabling the direct linking of morphological and gene expression data for advanced biological studies and drug development.
Implementation Method 1
an interdigital electrode is used to trap individual cells in the microwells
Implementation Method 2
capture oligonucleotides with unique barcode and identifier sequences capture mRNA
Data Source
AI summary
The present application provides a method and a system for integrating morphological characteristics and gene expression of individual cells. The method comprises the following steps: providing a microfluidic device, which comprises a microwell array and an interdigital electrode, and each microwell comprises a plurality of capture oligonucleotides; injecting cells into the microwells, capturing a single cell and recording morphological characteristics of the cell; lysing the cell so that the mRNA released by the cell is captured by the capture oligonucleotide; reverse transcribing the captured mRNA to obtain cDNA; performing a PCR amplification reaction on the cDNA to obtain a cDNA library and sequencing the cDNA library; reading the cell barcode sequence and the unique molecular identifier sequence according to sequencing results, and the morphological characteristics and gene expression of the cell in the microwell are integrated together.