Antibody Binding Analysis via Cell Partitioning and Nucleic Acid Barcoding
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Solution Overview
Problem
Current methods for assessing the binding specificity and affinity of antibodies to antigens are labor-intensive and low-throughput, often requiring assays like ELISA and SPR, which can be challenging for complex molecular targets.
Innovation Solution
A method involving the use of reporter oligonucleotides with antibody barcode sequences, which are contacted with cell populations expressing candidate antigens or epitopes. The method includes partitioning cells and generating barcoded nucleic acid molecules upon binding, allowing for the analysis of binding interactions through nucleic acid sequencing.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If traditional methods like ELISA and SPR are used to assess antibody binding, then binding specificity and affinity can be determined, but the process becomes labor-intensive and low-throughput
Solution Approach 1:
The method partitions a cell population into individual partitions (e.g., droplets or wells), with each partition containing a single cell or subset of cells. This segmentation enables parallel processing of multiple cells simultaneously, dramatically increasing throughput while maintaining precise binding measurements through partition-specific barcodes.
Solution Approach 2:
The invention introduces barcoded nucleic acid molecules as intermediaries to link antibody binding events to detectable signals. When an antibody binds to an antigen on a cell surface, it brings the reporter oligonucleotide into proximity with the cell's endogenous barcode, forming a hybrid molecule that serves as a measurable intermediary indicating the binding event.
2Measurement precision
If traditional assays are used for binding analysis, then binding data can be obtained, but the process requires multiple separate assays which increases time and complexity
Solution Approach 1:
The method combines multiple measurement functions into a single integrated assay. By using barcoded nucleic acid molecules that contain both cell identity information and antibody binding information, the system simultaneously performs cell identification, binding detection, and signal amplification in one experiment rather than requiring separate ELISA, flow cytometry, and sequencing assays.
Solution Approach 2:
The barcoded nucleic acid molecule serves multiple functions: it acts as a cell identifier through the endogenous barcode, a binding indicator through the reporter oligonucleotide, and a signal amplifier through nucleic acid sequencing. This multi-functionality eliminates the need for multiple specialized assays and reduces overall analysis time.
3Measurement precision
If complex molecular targets are analyzed using traditional methods, then binding can be assessed, but crystallization becomes challenging due to stability issues
Solution Approach 1:
The invention replaces the mechanical/physical approach of X-ray crystallography with a biochemical approach using nucleic acid hybridization and sequencing. Instead of requiring stable protein crystals for structural analysis, the method uses stable nucleic acid barcodes that can be sequenced from solution, avoiding crystallization challenges entirely while providing binding interaction data.
Data Source
AI summary
The present disclosure relates in some aspects to methods and compositions for assessing the target binding capacity of a binding molecule, such as an antigen binding molecule, e.g., a monoclonal antibody. In some embodiments, provided herein is a method of analyzing binding of a barcode-labeled antibody to a target antigen expressed by a cell, wherein cells expressing the candidate antigens are partitioned and wherein detection of the barcoded nucleic acid molecule is indicative of the binding between the antibody or antigen-binding fragment thereof and the candidate antigen or epitope of the cell.