PURIFICATION OF HUMAN ALPHA(2)-ANTIPLASMIN WITH CHICKEN IGY SPECIFIC TO ITS CARBOXY-TERMINAL PEPTIDE

alpha(2)-antiplasmin, a plasma glycoprotein of the serpin superfamily, is the primary physiological inhibitor of plasmin, the key enzyme in fibrin degradation. Previous purification methods utilize lengthy mult istep protocols with low yields or use monoclonal antibodies that are expensive or difficult to make. With a relatively small investment, a chicken was immunized with keyhole limpet hemocyanin-conjugated to alp ha(2)-antiplasmin C-terminal 26 residue synthetic peptide and the pept ide-specific antibody (IgY) was isolated from the egg yolks of hens us ing the peptide affinity column. Based on the interaction between this IgY and alpha(2)-antiplasmin, pure alpha(2)-antiplasmin was isolated from human plasma in two steps: (a) citrated plasma was precipitated w ith 15% PEG-8000 to remove the bulk of plasma proteins while retaining the majority of alpha(2)-antiplasmin activity; and (b) the alpha(2)-a ntiplasmin was affinity-purified from the supernatant using the IgY co lumn. Yields were typically 48% and the purity and authenticity of the alpha(2)-antiplasmin were verified by gel electrophoresis, Western Bl ot analysis, N-terminal sequence, and amino acid analysis.