Heparins from different species and tissues show similar levels of ATIII and HCII mediated anti-IIa activities. On fractionation, chains containing predominantly ATIII or HCII activities could not be separated. Oligosaccharide mapping demonstrates that the concentration of an oligosaccharide comprising a portion of heparin's ATIII binding site in a particular heparin fraction correlates with ATIII mediated anti-IIa activity, but does not correlate with HCII mediated anti-IIa activity. These results suggest that ATIII and HCII do not share a common binding site. Partial enzymatic depolymerization of heparin resulted in large oligosaccharides which could be purified and partially characterized. Although oligosaccharides of degree of polymerization (dp) 18 and 20 showed significant ATIII and HCII mediated anti-IIa activities no separation of these activities resulted. These data suggest however that a minimum chain length of dp18 was required for HCII mediated anti-IIa activity.