Microfluidic Epitope Mapping via Partitioned Barcoding
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Solution Overview
Problem
Current methods lack high-throughput and efficient means to characterize and identify antigen-binding molecules (ABMs) and their binding sites on target antigens, which is crucial for developing effective immunotherapies and responding to immune-escaping pathogen variants.
Innovation Solution
The method involves partitioning a reaction mixture containing cells expressing ABMs, a target antigen, and a first agent bound to a reporter oligonucleotide, generating barcoded nucleic acid molecules to distinguish cells binding at different epitopes, and using these barcodes to characterize ABMs with unique binding specificities.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If conventional methods are used to characterize antigen-binding molecules, then the process can identify binding sites, but the throughput is low and the process is inefficient
Solution Approach 1:
The patent segments the characterization process into distinct modular components: (i) partitioning the reaction mixture into multiple partitions, (ii) introducing different blocking agents to each partition, (iii) adding reporter oligonucleotides that bind to ABMs, and (iv) detecting binding patterns. This segmentation enables parallel processing of multiple ABMs simultaneously, dramatically increasing throughput while reducing the time required for complete epitope mapping.
2Productivity
If high-throughput methods are implemented, then productivity increases, but the device complexity and process complexity increase
Solution Approach 1:
The patent employs universal blocking agents that can bind to multiple different epitopes on target antigens, and universal reporter oligonucleotides that can detect multiple different ABMs. This multi-functionality allows a single standardized platform to characterize diverse ABM-antigen pairs without requiring specialized reagents for each combination, thereby increasing throughput while controlling system complexity through reagent standardization.
3Measurement precision
If detailed epitope mapping is performed, then measurement precision improves, but the time required and process complexity increase
Solution Approach 1:
The patent performs preliminary blocking of epitopes using blocking agents before introducing the ABMs and reporter oligonucleotides. This preliminary action prevents ABMs from binding to blocked epitopes, allowing precise identification of which epitopes are recognized by each ABM based on which reporter oligonucleotides bind. This approach achieves detailed epitope mapping precision while reducing time by eliminating the need for sequential individual epitope analysis.
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 approach enables rapid and efficient identification of ABMs with distinct epitope specificity, facilitating the development of new immunotherapies and combination therapies, particularly against emerging pathogens and cancers.
Implementation Method 1
a first agent, wherein the first agent binds the target antigen at a first target binding site
Implementation Method 2
the target antigen bound to the first agent is coupled to a first reporter oligonucleotide
Implementation Method 3
a reaction mixture, or a portion thereof, is partitioned into a plurality of partitions
Implementation Method 4
Barcoded nucleic acid molecules are generated
Implementation Method 5
The barcoded nucleic acid molecules include: (i) a first barcoded nucleic acid molecule comprising a sequence of the first reporter oligonucleotide
Data Source
AI summary
The present disclosure relates generally to compositions, methods, kits, partitions and systems for high-throughput microfluidic identification of antigen-binding molecules and antigen binding molecule epitope mapping. Particularly, the compositions, methods, kits, partitions and systems of the disclosure relate to the characterization of cell-expressed antigen-binding molecules (e.g., antibodies) produced by a population of cells, such as B cells, that may have different binding and epitope specificity to a target antigen of interest and/or with enhanced activity.