Scutoxin A and B represent two isoforms of a new toxic protein from the venom of the Australian tiger snake, Notechis scutatus scutatus. Both isoforms, of apparent mol. wt 13,000, are less basic than either notexin or notechis II-5. They both have similar i.v. LD50-values in mice of ca 0.006 micrograms/g, and phospholipase activities of about 136 mumoles of fatty acid released/min/mg at 37 degrees C when acting on phosphatidylcholine in the presence of Triton X-100. Toxicities of the scutoxins are the same as notexin and about seven times more potent than notechis II-5. ELISAs and western blot analyses indicate that the new toxins are immunologically similar to notexin and notechis II-5, with phospholipase activities falling between these latter two proteins. When crude venom is initially passed over a gel filtration column, each scutoxin isoform co-elutes in a different fraction with notexin. Gel filtration experiments using purified samples of notexin and scutoxin have failed to demonstrate any evidence for the formation of higher mol. wt protein complexes. Peptide mapping suggests the presence of five glutamate residues in one of the protein isoforms. These findings, together with the high toxicity and active phospholipase levels, demonstrate that the new proteins are not the previously reported non-toxic and enzymatically inactive notechis II-1. The combination of gel filtration on Sephacryl S-200 and cation-exchange chromatography used to isolate the scutoxins also permits recovery of notexin and notechis II-5 in high purity.