Summary
A technique is described to completely remove antithrombin III (AT) from small amounts
of human plasma by immunoaffinity chromatography on antibodies against human AT linked
to Sepharose 4B. The level of heparin cofactor II (HCII) was not affected by the immunoadsorption.
HCII activity was then determined by measuring the rate of human thrombin inhibition
by 3 ways: a) activation with heparin in AT-free plasma, b) activation with dermatan
sulfate in normal plasma and c) activation with dermatan sulfate in AT-free plasma.
The normal range of HCII varied between 0.7-1.5 U/ml, as compared to a normal plasma
pool containing by definition 1 U/ml. Highly significant correlations between assays
as obtained from 40 normal plasmas proved the suitability of the 3 assays, although
the progressive thrombin inhibition by AT, when not removed, contributed about one
fifth to the thrombin inhibition by HCII in the presence of dermatan sulfate. There
were also highly significant correlations between HCII activity and antigen, as determined
by rocket immunoelectrophoresis using specific antibodies against HCII.
Levels of HCII and AT were examined in 7 patients with hereditary AT deficiency and
7 patients with disseminated intravascular coagulation (DIC). In hereditary AT deficiency,
whereas the AT activity was reduced by half, levels of HCII activity and antigen were
in the normal range. In DIC, a parallel decrease of HCII and AT suggests that HCII
may participate in the inhibition of thrombin released during DIC and thus provides
an inhibitor reserve, once the AT level becomes subnormally low.
Keywords Heparin cofactor A - Antithrombin III - Disseminated intravascular coagulation - Antithrombin
III deficiency - Immunoaffinity chromatography
