Barcoded Antigen Display Sequencing for Rare B-Cell Specificity Mapping
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Solution Overview
Problem
Existing methods for linking antibody sequences to antigen specificity are fragmented, labor-intensive, and low-throughput, hindering the ability to detect rare clones and explore cross-reactivity in a scalable and integrated manner.
Innovation Solution
A method called dtLIBRA-Seq, which involves constructing cell-free barcoded antigen display libraries, allowing antigens to bind to B-cells, separating bound cells into single emulsions, and using unique barcodes and molecular identifiers to sequence and align antibody sequences with antigens, determining a LIBRA-seq score for each pair.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If traditional antibody discovery methods (single B-cell culture, hybridoma screening) are used, then antibody sequence can be obtained, but throughput is limited and labor-intensive
Solution Approach 1:
The patent combines antibody sequence acquisition and antigen-binding assay into a single integrated workflow using single-cell RNA sequencing. This merging eliminates the need for separate cloning and expression steps, achieving high-throughput processing while maintaining precise antibody sequence detection and functional specificity determination from the same cell.
Solution Approach 2:
The single-cell RNA sequencing approach serves multiple functions simultaneously: it captures antibody sequence information, determines antigen specificity, and enables high-throughput processing. This multi-functional methodology replaces multiple specialized techniques with a single comprehensive approach.
2Loss of information
If conventional techniques are used to detect antibody-antigen relationships, then some level of detection is achieved, but concurrent recovery of sequence and functional specificity from the same cell is rarely permitted
Solution Approach 1:
The patent merges sequence acquisition and functional assessment into a single single-cell RNA sequencing experiment. By extracting and sequencing mRNA from individual cells, the method simultaneously captures both the antibody sequence information and the antigen-binding functionality without requiring separate experimental steps or complex multi-stage workflows.
3Measurement precision
If traditional methods are employed, then antibody sequences can be identified, but the ability to detect rare clones and explore cross-reactivity in a scalable manner is hindered
Solution Approach 1:
The patent changes the key parameter from low-throughput single-cell culture to high-throughput single-cell RNA sequencing. This parameter change enables the processing of thousands of cells simultaneously, dramatically improving the ability to detect rare clones and explore cross-reactivity while maintaining the scalability needed for comprehensive immune repertoire analysis.
Data Source
AI summary
The present disclosure relates to methods for simultaneous detection of antigens and antibodies or host receptor proteins that specifically bind said antigens.