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Epitope mapping

Epitope mapping is performed to identify the epitope to which a monoclonal antibody binds.  

Conformational epitopes consist of residues that are scattered in the primary sequence but brought together through folding of the protein of interest. The determination of conformational epitopes is a major issue in the antibody field, the standard techniques for determining linear epitopes being ineffective.

Agro-Bio has therefore developed a robust method for identifying an antibody’s conformational epitopes using in situ proteolysis involving formation of an immobilized antigen-antibody complex, followed by mass spectrometry analysis.

Our method combines:

  • proteolytic digestion
  • isolation of the epitope peptides by affinity chromatography
  • final analysis of the epitope fragments by mass spectrometry.

Two approaches are possible:

  • the antigen/antibody complex is formed during affinity chromatography and the complex is digested within the affinity column
  • antigen digestion is performed before formation of the antigen/antibody complex inside the affinity column.

Identification of the sequence of interest is based on the fact that antigen/antibody complex formation protects the epitope from enzyme digestion. 

After affinity chromatography and digestion, the epitope fragments are eluted then sequenced by mass spectrometry: MALDI-TOF/TOF.

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