CreMap™ Epitope Mapping by Alanine Scanning Mutagenesis

Based on enriched experience and professional technical team, Creative Biolabs provides Alanine Scanning Mutagenesis library to help our clients performing B cell epitope mapping. And this method could identify the crucial amino acid residues of certain target peptides which contribute to the peptide’s functionality.

Alanine scanning library Fig 1. Alanine scanning library.

Alanine scanning mutagenesis technology is a useful and powerful approach to construct the peptide library (alanine scanning library) for identifying the B cell epitopes. Because of its inert methyl functional group, Alanine is often used on our platform to characterize the contribution of a specific amino-acid. Libraries involving other mutagenesis are also available when required. Each non-alanine residue of the target peptide will be sequentially substituted by one alanine at a time, which we could construct the mutation peptide library. A highly-specified site-directed mutagenesis is introduced in this crucial step. Then hundreds of plasmid clones from the mutation library will be expressed in mammalian cells. Subsequently, the corresponding change of the peptide-antibody binding activity will be measured. The antigen whose key amino-acid is replaced by the alanine loses or changes the binding ability to the specific antibody. Substitution of the key amino acid residue with alanine will lead to the mutation peptide’s failure of binding to antibodies or changes in the binding activity. Substitution with alanine residues eliminates side-chain interactions without altering the backbone conformation or introducing steric or electrostatic effects, so it is often the preferred choice for testing the contribution of specific side-chains while preserving native protein structure.

Workflow of CreMap™ Alanine Scanning Mutagenesis Mapping Service

Epitope Mapping by Alanine Scanning Mutagenesis

  • Each residue of the target protein is systematically substituted by alanine at a time through automated mutagenesis to construct the comprehensive mutation library.
  • Hundreds of plasmid clones from the mutation library are individually arrayed in the microplates, and used for high-throughput expression in mammalian cells.
  • Each mutation will be assayed in parallel for its contribution of binding to antibodies (or drugs, ligands). Based on that, the functional binding sites of the target peptide will be figured out.

Applications of CreMap™ Alanine Scanning Mutagenesis Mapping Service

CreMap™ alanine scanning mutagenesis mapping platform is a high-throughput strategy for mapping both linear and conformational antigen epitopes by evaluating the effects of point mutations across a target protein. And customs could obtain precise information about the functional binding sites of the target peptide at the level of amino acid. If you are interested in this service, please contact us for more information.

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