Shark IgNAR vNAR Binding Domains for Steric Hindrance Reduction
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Solution Overview
Problem
Current affinity capture assays face limitations due to steric hindrance and sub-optimal kinetics caused by the large size of intact immunoglobulins, which affects binding capacity and detection sensitivity, particularly for novel human antigens that do not bind well to existing VHH-based affinity resins.
Innovation Solution
The use of modified shark IgNAR-derived binding domains with a template vNAR sequence, immobilized on substrates via linkers, to create affinity resins and devices that enhance selectivity and binding capacity for a wide range of analytes, including novel human antigens.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If intact immunoglobulins are used as capture ligands, then binding capacity is achieved, but steric hindrance occurs due to large size affecting assay kinetics and binding capacity
Solution Approach 1:
The patent segments the immunoglobulin molecule into a single variable domain (vNAR) of approximately 100 residues, separating the antigen-binding function from the rest of the immunoglobulin structure. This segmentation eliminates steric hindrance while preserving binding capacity, as the compact vNAR domain can access epitopes that intact immunoglobulins cannot reach.
Solution Approach 2:
The patent extracts only the essential antigen-binding variable domain from the complete immunoglobulin molecule, discarding the constant regions and other domains that contribute to size but not to antigen recognition. This extraction creates a minimalistic binding unit that maintains affinity while reducing molecular weight and steric bulk.
2Weight of moving object
If VHH single domain antibodies are used, then smaller size is achieved, but binding affinity to novel human antigens is insufficient
Solution Approach 1:
The patent changes the structural parameters of the single domain antibody by adopting the vNAR framework from cartilaginous fish immunoglobulins, which has different complementarity determining region (CDR) configurations compared to camelid VHH. This parameter change in the protein scaffold enables access to novel human antigens that VHH cannot bind, while maintaining the small size advantage.
Solution Approach 2:
The patent creates a composite binding domain by combining the vNAR variable domain with engineered CDR regions that are specifically optimized for human antigen recognition. This composite structure integrates the stability and small size of single domain antibodies with enhanced affinity for human-specific epitopes.
3Productivity
If affinity capture assays use existing ligands, then detection capability is achieved, but sensitivity is limited for novel human antigens
Solution Approach 1:
The patent changes the binding parameters by introducing vNAR domains with altered CDR loops and framework regions that are evolutionarily distinct from mammalian antibodies. This parameter change enables the detection of novel human antigens with higher sensitivity, as the vNAR can recognize epitopes that are not accessible to conventional antibody formats.
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
The modified IgNAR-based affinity resins and devices offer improved selectivity and binding affinity, enabling robust and sensitive detection and enrichment of analytes, including novel human antigens, with high capacity and stability, suitable for various biological samples and applications.
Implementation Method 1
modified cartilaginous fish-derived immunoglobulin-like molecules, IgNARs, having desirable functions, such as binding affinity to one or more targets
Data Source
AI summary
The present disclosure relates to protein arrays and methods of using the same for the detection, quantification and characterization of biomolecules that specifically bind to the array among various other biomolecules in a biological sample. Specifically, the present disclosure relates to protein arrays that include a plurality of immunoglobulin molecules derived from shark single-domain heavy chain antibody lacking light-chains but including at least one variable antigen-binding domain with a binding site for an antigen. The immunoglobulin molecules are immobilized on a substrate via a linker. Further encompassed herein are diagnostic devices and kits, comprising the protein array and methods of using same.