Compositions and methods for identifying nanobodies and nanobody affinities

The method addresses the challenges of identifying antigen-specific nanobody repertoires by using mass spectrometry and antigen-specific affinity chromatography to select sequences with sufficient fragmentation coverage, achieving precise and sensitive Nb analysis.

JP7765090B2Active Publication Date: 2025-11-06UNIV OF PITTSBURGH OF THE COMMONWEALTH SYST OF HIGHER EDUCATION
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Patent Information

Application Number
JP2022566362
Authority / Receiving Office
JP · JP
Patent Type
Patents
Current Assignee / Owner
Priority Date
2020-05-01
Filing Date
2021-04-29
Publication Date
2025-11-06
Estimated Expiration
2041-04-29

AI Technical Summary

Technical Problem

Current methods for identifying and analyzing antigen-specific nanobody (Nb) repertoires face challenges due to the high diversity and dynamic range of circulating antibodies, large sequence databases with conserved frameworks, and limitations in efficient protocols and informatics for accurate quantification and classification, making it difficult to reliably identify and quantify Nb sequences.

Method used

A method involving obtaining a blood sample from a camelid immunized with an antigen, creating a cDNA library, isolating Nbs with trypsin or chymotrypsin, performing mass spectrometry analysis, and selecting sequences with sufficient fragmentation coverage to reduce false positives, combined with antigen-specific affinity chromatography and deep learning models for classification.

Benefits of technology

This approach enables accurate and sensitive identification of Nanobody sequences with reduced false positives, allowing for reliable analysis and classification of antigen-specific Nb proteomes, enhancing the precision of Nb discovery and characterization.

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Abstract

Provided herein are methods for identifying a set of Nanobody amino acid sequences (CDR3, CDR2 and / or CDR1 sequences) for complementarity determining regions (CDRs) 3, 2, and / or 1, where the reduced CDR3, CDR2 and / or CDR1 sequences are false positives compared to a control, methods for determining the antigen affinity of Nanobody peptide sequences, and related methods for training deep learning models.
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Citation Information

Patent Citations

  • Method and system for screening nanobody

    US20170226566A1

  • Compositions and methods for rapid production of versatile nanobody repertoires

    US20190391159A1