Preserving Antibody Chain Pairing in Nucleic Acid Libraries
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Solution Overview
Problem
Current methods for identifying antibodies are limited by the inability to effectively pair heavy and light chains from memory B-cells or plasma cells, leading to underrepresentation of useful antibody sequences, particularly those relevant to immune responses against infectious diseases, cancer, autoimmune diseases, and neurodegenerative diseases.
Innovation Solution
The development of nucleic acid libraries that preserve the in vivo multimeric associations of immune binding proteins, allowing for the enrichment of functional pairs of light and heavy chains from individuals who have mounted an immune response, using techniques such as overlap extension primers and barcode-based sequencing to maintain the pairing of polypeptide chains.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If conventional antibody library methods are used to amplify nucleic acids from B cells, then antibody sequences can be obtained, but the pairing of heavy and light chains is lost leading to underrepresentation of useful antibodies
Solution Approach 1:
The patent embeds both heavy and light chain nucleic acid sequences within a single vector structure that maintains their in vivo pairing relationship. This nested organization ensures that when the vector is amplified and expressed, the heavy and light chains remain correctly paired throughout the library generation process, solving the loss of pairing accuracy while maintaining sequence diversity
Solution Approach 2:
The patent combines the heavy chain and light chain nucleic acid sequences into a single integrated construct or vector system. By merging these previously separate amplification processes into one unified approach, the method preserves the natural pairing of chains while capturing diverse antibody sequences from B cells
2Quantity of substance
If antibody libraries are constructed from experimental captures of B cell sequences, then some antibody sequences are obtained, but useful paired light and heavy chains from successful immune responses are completely lacking or under-represented
Solution Approach 1:
The patent performs preliminary enrichment of B cells that have undergone successful immune responses before library construction. By pre-selecting activated B cells, memory B cells, or plasma cells that have encountered antigens in vivo, the method ensures that the captured antibody sequences represent functionally useful pairs from successful immune responses rather than random or naive sequences
3Productivity
If conventional library methods amplify nucleic acids from B cells, then antibody sequences are generated, but the in vivo multimeric associations of immune polypeptide chains are not preserved
Solution Approach 1:
The patent nests multiple polypeptide chain sequences (heavy and light chains) within a single stable vector structure that maintains their native associations. This nested organization protects the multimeric relationships during amplification and storage, preserving the in vivo composition stability while enabling efficient library generation
Solution Approach 2:
The patent creates accurate copies of the in vivo multimeric associations by using PCR or other amplification methods that replicate the paired heavy and light chain sequences together. These copied pairing relationships are then maintained through vector-based expression systems, ensuring that the amplified library preserves the original compositional stability
Data Source
AI summary
Described herein are immune binding proteins and method for obtaining immune binding proteins from genomic or other sources. Also described herein are nucleic acids encoding the immune binding proteins in which the natural multimeric association of chains is maintained in the nucleic acids and the immune binding proteins made therefrom. For example, nucleic acids encoding antibodies that are amplified from a B-cell using the methods described herein maintain the natural pairing of heavy and light chains from the B-cell. This maintenance of pairing (or multimerization) produces libraries and/or repertoires of immune binding proteins that are enriched for useful binding molecules.