Identification and characterization of human eosinophil cationic protein by an epitope-specific antibody

The eosinophil cationic protein (ECP) is a basic secretion proteininvolved in the immune response system. ECP levels in biological fluidsare an indicator of eosinophil-specific activation and degranulationand are currently used for the clinical monitoring and diagnosis ofinflammatory disorders. A polyclonal epitope-specific antibody has beenobtained by immunizing rabbits with a conjugated synthetic peptide. Asequence corresponding to a large exposed loop in the human ECPthree-dimensional structure (D115–Y122) was selected as a putativeantigenic epitope. The antibody was purified on an affinity columnusing recombinant ECP (rECP) as antigen. The antibody (D112–P123 Ab)specifically recognizes rECP and its native glycosylated andnonglycosylated forms in plasma, granulocytes, and sputum. The antibodydetects as little as 1 ng of rECP, can be used both in reducing andnonreducing conditions, and does not cross-react with the highlyhomologous eosinophil-derived neurotoxin or other proteins of thepancreatic ribonuclease superfamily.