Bioassay-directed purification of an acidic phospholipase A2 from Agkistrodon halys pallas venom

Relying on ex vivo and in vitro platelet anti-aggregation assays, a tail bleeding time assay, and an anti-thrombotic assay, we have purified the fraction of venom from Agkistrodon halys pallas which, in all of these assays, is the most active. There were two major steps in the purification: gel filtration chromatography on Sephadex G-100, and ion exchange chromatography on DEAE Sephadex A-50. Sequencing of the most active fraction by mass spectrometry revealed that it is a known acidic phospholipase A2. Prior expectations by others about the in vivo anti-thrombotic activity of this enzyme are confirmed.