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SummaryA relatively low heparin cofactor activity (0.60 U/ml) was observed in a patient with recurrent superficial thrombophlebitis of the left leg. However, the antigen concentration was in the normal range (1.04 U/ml) and the progressive antithrombin activity was normal. The crossed immunoelectrophoresis in presence of heparin in agarose gel separated the patient's AT-III antigen in 2 fractions with different mobilities. The patient's AT-III was purified for further characterization. The last step of the purification procedure, a heparin-agarose chromatography, led to a separation and a purification of 2 AT-III fractions with different heparin affinities: an abnormal AT-III with reduced heparin affinity and a normal AT-III with a heparin affinity similar to that of AT-III isolated from normal plasmas. Abnormal and normal AT-III share several identical properties as molecular weight, ability to form complexes with thrombin and progressive antithrombin activity.

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SummaryAssuming 1 U/ml in titrated plasma, the VIII: CAg concentration was found 1.66 U/ml in EDTA-plasma, 1.09 U/ml in heparinized plasma and 0.67 U/ml in serum. Addition of 10 mmol/1 EDTA to titrated and heparinized plasmas increased VIII: CAg 1.5fold. There was no increase of VIII: CAg in serum. Gel filtration of plasmas on different anticoagulants showed an elution of VIII: CAg in the void volume Vo and in the later fractions. The VIII: CAg amount detected in the internal volume increased following the series heparin < citrate < EDTA. Serum VIII: CAg was eluted at 2.2 Vo. Presence of EDTA in the elution buffer or incubation of plasma with EDTA prior to chromatography caused a displacement of practically all VIII: CAg amount in the internal volume with a peak at 2.2-2.3 Vo. VIIIR: Ag was exclusively detected in the void volume.Removal of divalent cation by chelation likely exposes more antigenic determinants of VIII: CAg, which are otherwise masked by steric hindrance due to VIIIR: Ag in citrated and heparinized milieu. Moreover gel filtration of plasma in the presence of EDTA completely dissociates VIII: CAg from VIIIR :Ag. The VIII: CAg fragment, having an estimated molecular weight of 70,000, might also be present in serum.

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