Epitope Barcode Single-Cell Detection System
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Solution Overview
Problem
Current methods for monitoring gene expression profiles in individual cells are often inefficient, requiring large biological samples and failing to provide cell-specific information, especially when analyzing complex samples or detecting specific variants of proteins and nucleic acid sequences.
Innovation Solution
The use of oligonucleotide proximity probes with epitope-specific barcode sequences and bridge oligonucleotides to detect and quantify target nucleic acid sequences in single cells, allowing for the creation of unique cell origination barcodes that identify individual cells within a mixture.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional methods are used to monitor gene expression profiles, then detection can be performed, but large amounts of biological sample are required and cell-specific information is lost
Solution Approach 1:
The method segments the detection process by assigning unique barcode sequences to individual cells through oligonucleotide proximity probes that bind to cell-specific targets. Each cell's gene expression profile is monitored separately with its own barcode identifier, enabling single-cell resolution without requiring large sample pools.
Solution Approach 2:
Barcodes serve as intermediary molecules that link individual cells to their gene expression profiles. The barcodes are introduced into cells and used to tag and identify specific cells during sequencing, allowing cell-specific information to be retrieved from bulk sequencing data.
2Adaptability or versatility
If multiplexed measurements are performed to detect multiple target molecules, then comprehensive analysis is achieved, but the complexity of analysis increases significantly
Solution Approach 1:
The barcode system provides a universal solution for multiplexed measurements by using a common barcode structure that can identify multiple different target molecules simultaneously. The same barcode tagging approach works for various nucleic acid targets, simplifying the analysis framework while maintaining versatility.
Solution Approach 2:
The method adds a barcode dimension to the detection space, transforming complex multiplexed measurements into a more manageable format. By encoding cell identity and target molecule information in barcode sequences, the system separates the complexity of multiplexing from the analysis burden through sequence-based identification.
3Loss of information
If bulk analysis is performed on complex samples, then processing is simplified, but cell-specific information regarding target molecules is lost
Solution Approach 1:
The method creates a copy of cell identity information in the form of barcodes that are introduced into individual cells. These barcode copies allow bulk processing of samples while preserving cell-specific information, as each cell's barcode sequence serves as a permanent identifier that can be retrieved and traced back to the original cell.
4Measurement precision
If single-cell analysis is performed to retain cell-specific information, then precise cell identification is achieved, but the difficulty of detecting and measuring increases
Solution Approach 1:
Barcodes act as intermediary molecules that simplify single-cell detection by providing a direct molecular link between the cell and its identification code. The barcodes are introduced into cells and can be amplified and sequenced along with the target molecules, making single-cell analysis as straightforward as bulk 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 accurate and sensitive detection and quantification of target molecules in individual cells, retaining cell-specific information and facilitating the identification of specific sub-populations within complex cell populations.
Implementation Method 1
a first oligonucleotide proximity probe comprising an epitope specific barcode sequence and a first target recognition sequence that is capable of hybridizing to a first segment of the target nucleic acid sequence
Implementation Method 2
providing a bridge oligonucleotide that comprises two probe recognition sequences, wherein the first probe recognition sequence is capable of hybridizing to a segment of the first oligonucleotide proximity probe, and the second probe recognition sequence is capable of hybridizing to a segment of the second oligonucleotide proximity probe, thereby creating a target specific probe complex
Data Source
AI summary
A method for identifying a sub-population within a mixed population of cells is disclosed. The method involves contacting the mixed population of cells with at least one unique binding agent, wherein the at least one unique binding agent is designed to bind to a target molecule present in the sub-population, and wherein the at least one unique binding agent is attached to an epitope specific barcode that represents the identity of the target molecule. The method further involves sequentially attaching two or more assayable polymer subunits to the epitope specific barcode to create unique cell origination barcodes that represent the identities of individual cells to which the at least one unique binding agent has bound; and decoding the epitope specific barcode and cell origination barcodes, thereby identifying the sub-population within the mixed population of cells.