miRNA-Linked mRNA Marker for Live Cell Type Identification
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Solution Overview
Problem
Current methods for distinguishing cell types are limited in accuracy and applicability, particularly in live cells, as they often require cell immobilization or membrane permeation, and are not suitable for refining classifications due to surface receptor limitations and limited detectable combinations.
Innovation Solution
A method using mRNA with a specific marker gene linked to target miRNA sequences to differentiate cell types by translating the marker gene, allowing for live cell analysis and high-resolution classification using flow cytometry or imaging analyzers.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If antibody-based detection is used to identify cell surface ligands, then cell type identification can be performed, but cell immobilization and membrane permeation are required which prevents fractionation of living cells
Solution Approach 1:
The patent introduces an intermediary system consisting of cell-penetrating peptides conjugated to detection molecules. These peptides mediate the delivery of detection agents into living cells without requiring cell immobilization or membrane permeation procedures, thereby enabling identification of intracellular markers in living cells while maintaining cell viability and functionality
2Measurement precision
If qualitative classification using antibody-based ligand detection is employed, then cell types can be distinguished, but detectable combinations are limited making refined classification difficult
Solution Approach 1:
The patent segments the classification process into multiple independent detection dimensions by utilizing different cell-penetrating peptides with distinct sequences and properties. Each peptide can target different intracellular markers, allowing for combinatorial detection strategies that enable refined classification of cell types through multiple independent measurement channels rather than relying on limited surface receptor combinations
3Measurement precision
If microarray or next-generation sequencing is used for quantitative cell classification, then cells can be quantitatively classified based on multiple molecules, but the measured cells are destroyed making live cell measurement impossible
Solution Approach 1:
The patent employs disposable cell-penetrating peptide conjugates that can be introduced into living cells for transient detection. These peptide-based detection agents are designed to be non-integrative and temporary, allowing quantitative measurement of intracellular markers without permanent genetic modification or cell destruction, thus enabling live cell quantitative classification unlike destructive methods such as microarray or sequencing
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 high-accuracy, live-cell differentiation of cell types with reduced risk of genetic damage and improved detection precision, suitable for medical applications like pluripotent stem cell differentiation, even with slight miRNA expression differences.
Implementation Method 1
introducing mRNA comprising a first marker gene operably linked to the target sequence of miRNA used as an indicator into a cell group
Data Source
AI summary
A method for distinguishing living cells in a living state with high accuracy. A method for distinguishing a desired cell type from a cell group comprising two or more types of cells, using the expression of miRNA as an indicator, wherein the method comprises the following steps:(1) a step of introducing mRNA comprising a marker gene operably linked to the target sequence of miRNA used as an indicator into a cell group; and(2) a step of distinguishing a cell type, using the translation level of the marker gene as an indicator.