Ditazole (4,5-diphenyl-2-bis-(2-hydroxyethyl)-aminoxazol) has been shown to be a strong in vitro inhibitor of human platelet aggregation brought about by release reaction inducers; in contrast, it did not significantly affect primary ADP-induced aggregation. Ditazole strongly inhibited the release of platelet-bound ¹⁴C-serotonin under the influence of Thrombofax, whereas it did not interfere with the transport and storage of serotonin in nonstimulated platelets. The effect of ditazole was not potentiated by acetyl-salicylic acid. Ditazole also inhibited ADP-reptilase clot retraction and modified thrombin-induced clot formation. The inhibition of platelet aggregation exerted by ditazole in plasma could be removed following gel filtration of platelets on Sepharose 2-B gel. This would indicate that ditazole does not act on platelets by a ‘hit and run’ mechanism.

Since fibrinogen has itself an antithrombin activity, defibrinogenation of the sample is a prerequisite for AT-III determination. Five different defibrinogenation procedures were tested: heat precipitation for either 3 or 10 minutes, ristocetin precipitation, clotting with either Reptilase-R or Botropase-R. AT-III was measured as biological activity according to Howie et al. (Br. J. Haemat. 25, 101, 1973) and assayed immunologically by radial immunodiffusion. Defibrinogenation was complete with all the procedures used (< 16 μg/ml F. R.-antigen being left). In the samples defibrinogenated by ristocetin, the biological assay could not be performed, since some ristocetin remained in the supernatant and provoked a rapid precipitation of the substrate fibrinogen used for the assay. The possibility that Botropase-R or Reptilase-R had themselves some antithrombin activity was preliminarly ruled out. In samples definbriogenated by heating (3 min), Reptilase-R or Botropase-R the same biological AT-III activity was found; in addition, no differences in immunological reactivity of the protein were observed as compared with the corresponding-plasma before defibrinogenation. In contrast, a statistically significant (P < 0.001) reduction of both biological and immunological AT-III activity was found in the samples defibrinogenated by heating (10 min), as compared with the other 3 procedures. These results indicate that Reptilase-R and Boropase-R are suitable reagents for defibrinogenation of plasma prior to AT-III assay; heat precipitation can also be used, provided the duration of the incubation period is accurately controlled.(Supported by Grant N. 73.00400.04 of the Italian Research Council (C. N. R.).)