Laboratory diagnosis of hereditary thrombophilia

Genetic defects of antithrombin (AT) or one of the components of the protei n C pathway are associated with hereditary thrombophilia. Laboratory assays are currently available to diagnose and type hereditary thrombophilia due to deficiency or dysfunction of one of the anticoagulant factors antithromb in (AT), protein C (PC) and protein S (PS), and APC resistance without the need of DNA analysis. There are no functional tests for the prothrombin mut ant G20210A and thrombomodulin mutations, which can be diagnosed by a PCR-b ased test or by gene analysis, respectively, Hereditary AT deficiency is cl assified in a quantitative type I and three functional type II deficiencies affecting the reactive site (RS), heparin binding site (HBS), or pleiomorp hic site of the AT protein, All four types of hereditary AT deficiencies ca n be diagnosed by a heparin cofactor assay and one immune assay in combinat ion with crossed immunoelectrophoresis of the AT protein. The combination o f an enzyme-linked immunoadsorbent assay (ELISA) and a functional Protac-AP TT-based assay for PC will detect quantitative type I and dysfunctional typ e II PC deficiencies. There is a significant overlap in PC antigen and func tional levels between heterozygotes of PC deficiency and normals leaving a gray zone of uncertainty in differentiating congenital PC deficiency and no rmal individuals, Accurate diagnosis of hereditary PS deficiency should be a combination of tests aimed to measure free PS activity and antigen and to tal PS antigen levels. APTT-, Xa-, and RVVT-based APC-resistance tests, whe n test plasmas are diluted in factor V deficient plasma, have increased in sensitivity and specificity to 100% for the discrimination of normal indivi duals from heterozygotes and homozygotes for factor V Leiden. The RVVT-base d APC-resistance test provides better separation of factor V Leiden and nor mals in the various clinical settings, lupus anticoagulant in particular. T he modified APC-resistance tests also claim a separation between heterozygo tes and homozygotes for factor V Leiden in the normal population, asymptoma tic subjects, and thrombosis patients. Below a certain cut-off level, a min or overlap of normalized APC ratios between heterozygotes and homozygotes f or factor V Leiden of thrombosis patients has been shown in one study, whic h still points to the need to perform the more time consuming and expensive DNA test to identify heterozygotes from the more clinically significant ho mozygotes, The prothrombin-based APC-resistance test, which measures thromb in activated factor Va in highly diluted test plasma, appears to be the mos t sensitive and specific of all APC-resistance tests and separates normal i ndividuals from heterozygotes and heterozygotes from homozygotes for factor V Leiden without the need of confirmation by a DNA test.