Barcoded Antigen Screening for Ligand-Blocking Antibody Potency
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Solution Overview
Problem
Current methods for characterizing antigen-specific B cells are low throughput, limiting the ability to screen individual B cells against multiple antigens simultaneously, and there is a need for high-throughput systems to detect ligand-blocking antibodies and determine antibody potency.
Innovation Solution
A method involving labeling antigens and ligands with unique barcodes, mixing with B-cells, separating into single cell emulsions, preparing cDNA libraries, and performing PCR amplification to generate amplicons with unique molecular identifiers (UMIs) for sequencing and alignment, determining LIBRA-seq scores to identify blocking interactions.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If traditional single-cell sorting or B cell culture methods are used to characterize antigen-specific B cells, then measurement precision can be maintained, but productivity is severely limited to low throughput
Solution Approach 1:
The method segments the B cell population into individual single cells within droplets, with each droplet containing a unique cell barcode. This segmentation enables parallel processing of thousands of individual B cells simultaneously through high-throughput sequencing, resolving the contradiction between maintaining single-cell resolution (measurement precision) and achieving high throughput (productivity).
Solution Approach 2:
The invention creates digital copies of B cell information through barcoded antigens and ligands that are sequenced alongside the B cell receptor sequences. This copying approach allows the physical state of B cell-antigen-ligand interactions to be captured and analyzed computationally, enabling high-throughput analysis without compromising the precision of individual cell characterization.
2Productivity
If high-throughput screening methods are implemented to increase productivity, then time loss is reduced, but device complexity increases due to multiple barcoding and sequencing requirements
Solution Approach 1:
The barcoded antigen and ligand molecules serve multiple functions simultaneously: they identify the specific antigen and ligand bound, provide unique cell barcodes for tracking individual B cells, and enable quantitative measurement of binding interactions. This multi-functionality reduces the need for separate systems for each measurement type, managing device complexity while maintaining high throughput.
Solution Approach 2:
The method merges the antigen screening, ligand blocking detection, and B cell receptor sequencing into a single integrated high-throughput workflow. By combining these functions into one assay using barcoded molecules and single-cell droplet technology, the system achieves high productivity without proportionally increasing device complexity.
3Productivity
If multiple antigens are screened against individual B cells simultaneously to increase productivity, then time loss is reduced, but measurement precision may be compromised by cross-contamination or signal interference
Solution Approach 1:
Each B cell is isolated in its own droplet with unique barcodes, physically segmenting the screening process to prevent cross-contamination between cells. The barcoded antigens and ligands are also segmented into discrete molecular identities, allowing precise tracking of which antigen and ligand bound to which B cell, maintaining measurement precision while enabling high-throughput parallel screening.
Solution Approach 2:
The sequencing data provides feedback that links specific B cell receptor sequences to specific antigen and ligand barcodes. This feedback mechanism allows computational analysis to accurately determine which antigens bound to which B cells and whether ligand blocking occurred, maintaining measurement precision even when screening multiple antigens simultaneously across thousands of cells.
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 high-throughput detection of ligand-blocking antibodies and determination of antibody potency, allowing for efficient screening and identification of potent antibodies with specific binding properties.
Implementation Method 1
allowing the plurality of barcode-labeled antigens to bind to the population of B-cells
Implementation Method 2
labeling one or more ligands to one or more antigens in the plurality of antigens with unique ligand barcodes
Data Source
AI summary
The present disclosure relates to high-throughput systems and methods for the detection of ligand-blocking antibodies and for determining antibody potency.