Barcoded Antibody Libraries via Ribosome Display
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Solution Overview
Problem
Current methods for producing scalable and cost-effective antibody libraries for detecting proteins are laborious and expensive, particularly when targeting multiple human protein targets simultaneously, as they rely on barcoding and pooling individual antibodies, which is inefficient compared to RNA or DNA sequence detection methods.
Innovation Solution
The development of cell-free platforms using distinct antibodies or antigen binding fragments, such as nanobodies, that are naturally barcoded with their encoding RNA by ribosome display, enabling parallel selection and large-scale production of libraries that can bind hundreds to thousands of cell surface proteins, with methods involving CRISPR/Cas9 for target identification and single-cell sequencing.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If barcoding and pooling of individual antibodies is used for detecting multiple protein targets, then detection capability is improved, but labor and cost increase significantly
Solution Approach 1:
The patent combines multiple antibody detection functions into a single pooled library where multiple barcoded antibodies coexist in one population. This merging approach eliminates the need for separate processing of individual antibodies, reducing labor while maintaining the ability to detect multiple protein targets simultaneously through high-throughput sequencing of the pooled library.
Solution Approach 2:
The pooled antibody library serves multiple detection functions simultaneously. A single library population can detect numerous different protein targets, making the system universally applicable across many detection scenarios without requiring separate antibody preparations for each target, thereby reducing both labor and cost.
2Adaptability or versatility
If large-scale antibody libraries are produced for hundreds to thousands of protein targets, then detection coverage is improved, but production complexity increases
Solution Approach 1:
The patent segments the large-scale library production into manageable components: individual antibodies are barcoded with unique identifiers, then pooled together. This segmentation allows the complex task of producing libraries for thousands of targets to be broken down into standardized units that can be efficiently managed and processed through automated high-throughput sequencing workflows.
Solution Approach 2:
The patent introduces barcodes as an intermediary element that links antibodies to their encoding information. This intermediary system enables automated tracking and identification of antibodies within the pooled library, reducing production complexity by replacing manual tracking with automated barcode reading and sequencing technologies.
3Measurement precision
If individual antibody barcoding is performed for each antibody, then antibody identification is improved, but time and resource consumption increase
Solution Approach 1:
The patent merges the identification process for multiple antibodies into a single high-throughput sequencing operation. Instead of identifying each antibody separately, the pooled library with barcoded antibodies is processed together, maintaining precise antibody identification through unique barcodes while dramatically reducing the time and resources required by performing all identifications in parallel.
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 allows for rapid synthesis and identification of large-scale libraries of barcoded antibodies, enabling high-dimensional cell cytometry, spatially indexed sequencing, and targeted proteomics, while systematically measuring off-target binding and mapping antibodies to specific proteins, facilitating the detection and perturbation of conventional and unconventional targets across various platforms.
Implementation Method 1
The antibodies or antigen binding fragments are naturally barcoded with their encoding RNA by ribosome display
Implementation Method 2
incubating antibodies or antigen binding fragments (e.g., RNA-barcoded nanobodies) with cells expressing a library of clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 single guide RNAs (sgRNAs) targeting genes
Implementation Method 3
Single-cell sequencing of antibody or antigen binding fragment (e.g., nanobody) sequences and sgRNA barcodes allows identification of sgRNAs associated with reduced binding of specific antibodies or antigen binding fragments
Data Source
AI summary
The present invention discloses high-throughput methods for the creation of antibodies or antigen-binding fragments that can bind to single or multiple targets. Also disclosed are methods for using one or more antibodies or antigen-binding fragments to detect cognate binding partners in various types of samples.