Summary
Rabbit polyclonal anti-protein S serum was fractionated with immobilized human protein S to establish solid-phase immunoradiometric assays recognizing Ca(II)-dependent and NonCa(II)-dependent epitopes of human protein S. The two assays were specific for PS:Ca(II)Ag and PS:NonCa(II)Ag and highly sensitive with a lower limit of detection of about 2.5 ng/ml.
PS:Ca(II)Ag and PS:NonCa(II)Ag levels were measured in immunopurified protein S, thrombin-modified protein S and chy-motrypsin-cleaved protein S. Only in chymotrypsin-cleaved protein S an important discrepancy between the two antigen levels was observed.
Ranges for the concentration of PS:Ca(II)Ag and PS:NonCa(II)Ag and their ratio were established in plasma of healthy individuals (0.92 ± 0.13 U/ml, 0.98 ± 0.21 U/ml, 0.96 ± 0.17, respectively). In a group of patients using oral anticoagulant therapy the ratio PS:Ca(II) Ag/PS : NonCa(II)Ag decreased at increasing intensity of anticoagulation suggesting the presence of sub- and noncarboxylated protein S molecules.
In plasma of patients with a hereditary type I protein S deficiency PS:Ca(II)Ag and PS:NonCa(II) Ag were reduced to the same extent: mean ratio 1.02 ± 0.12 in the group not on oral anticoagulant treatment and 0.94 ± 0.10 in the group on oral anticoagulant therapy. Analysis of patients with a history of unexplained thrombo-embolic disease did not reveal individual patients with a PS:Ca(II)Ag/PS:NonCa(II)Ag ratio below the lower limit of the normal range (mean ratio 1.05 ± 0.17), suggesting that the frequency of genetic protein S variants with defects in the Ca(II)-stabilized conformation is very low.
Keywords
Protein S antibodies - Oral anticoagulation - Calcium - Immunoradiometric assay - Thrombotic disease
