SNAIL Oligonucleotide Primers for Multiplexed Single-Cell Epitope Detection
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Solution Overview
Problem
Current methods for detecting and quantifying target molecules in cells require significant biological samples and fail to provide cell-specific information, with limited multiplexed protein measurement technologies due to challenges in protein samples.
Innovation Solution
The method involves using a pair of SNAIL oligonucleotide primers, including a Splint Primer Oligonucleotide (SPO) and a Padlock Oligonucleotide (PO), which form a circle upon binding, allowing for rolling circle amplification and the generation of a cell-specific barcode through the addition of assayable polymer subunits (APS) during split pool synthesis.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional detection methods are used to detect target molecules in cells, then detection can be performed, but significant amounts of biological samples are required and cell-specific information is lost
Solution Approach 1:
The method segments the detection process by first isolating individual cells through microfluidic droplet encapsulation, then performing target molecule detection within each isolated compartment. This segmentation allows detection of target molecules in minimal sample amounts while preserving cell-specific information, as each droplet contains a single cell and its detection products are physically separated from other cells.
2Adaptability or versatility
If multiplexed protein measurement technologies are used, then multiple target molecules can be measured, but the technology is limited due to challenges inherent in protein samples
Solution Approach 1:
The method replaces direct protein detection with nucleic acid-based detection. Since nucleic acids are easier to amplify and detect with high specificity, the invention uses reverse transcription PCR or other nucleic acid amplification techniques to detect protein expression levels indirectly through their mRNA transcripts. This substitution enables multiplexed measurement of multiple protein targets while avoiding the technical challenges of direct protein analysis.
3Loss of information
If conventional detection methods are used, then target molecules can be detected, but cell-specific information regarding target molecules is not retained
Solution Approach 1:
The method implements a nested structure where cell-specific information is encapsulated within droplets, which contain the cell and its detection products. The droplet acts as a physical container that preserves the association between the target molecule detection signal and its cell of origin. This nested architecture allows simultaneous preservation of cell identity and detection accuracy, as the detection products remain physically linked to their source cell throughout the analysis process.
Data Source
AI summary
The invention provides methods, compositions, kits and devices for the detection of target molecules. In some embodiments, the invention allows for multiplexed target molecule detection.