Evaluation of Anticoagulants for Serologic Assays of Cholera Vaccination

Blood collected with an anticoagulant is beneficial for simultaneous evaluation of immune cells and humoral components such as antibodies. However, it is critical that the anticoagulant does not affect quantitative and qualitative analyses of antibodies. In the present study, we examined the potential interference of the widely used anticoagulants heparin, EDTA, and acid citrate dextrose (ACD) on vibriocidal antibody activities and Vibrio cholerae lipopolysaccharide (LPS)-specific IgG, IgM, and IgA levels in the plasma and sera obtained from cholera patients or vaccinees. Serum vibriocidal antibody titer was inhibited in the presence of EDTA or ACD but not in the presence of heparin. Moreover, 100% (8/8) of the vibriocidal antibody titers of plasma samples obtained from the vaccinees in tubes containing heparin were identical to the titers of matched sera when compared with 37.5% (3/8) and 50% (4/8) of the plasma samples prepared with EDTA and ACD, respectively. Among LPS-specific Igs, the Pearson correlation coefficient (r) for IgA in serum and plasma was low (r = 0.716), and the coefficients for IgG and IgM were relatively high (r = 0.997 and r = 0.945, respectively) in heparinized plasma samples compared with the coefficient for IgG and IgM of EDTA- and ACD-treated plasma. Our results suggest that heparin is an appropriate anticoagulant for the collection of blood when measuring vibriocidal activities and antibody levels in plasma samples.     

Anticoagulants for avian and reptilian blood: Heparin and EDTA

Summary The effectiveness of EDTA (dipotassium and disodium salts) and heparin as anticoagulants for the blood of puffadders and pigeons has been investigated. In the case of the snakes all three substances produced haemolysis after variable time intervals, and increases in cell volume were observed. In the case of the pigcons, samples stored at 4° C showed lower haematocrit values and haemolysed sooner than samples stored at room temperature.     

Effect of anti-beta2glycoprotein I Lupus Anticoagulants on fibrin polymerization and fibrinolysis

Anti-beta2-Glycoprotein I (beta2GPI) autoantibodies are the prominent laboratory feature of Hughes syndrome. By prolonging some coagulation tests in the presence of exogenous phospholipids (PL), they behave as classical Lupus Anticoagulants (LA). We investigated the effect of 3 affinity-purified anti-beta2GPI IgG preparations from patients with Hughes syndrome on fibrin polymerization and fibrinolysis of normal plasma, measured by comparing the optical densities of assay mixtures in the presence of the autoantibodies or normal IgG. The presence of anti-beta2GPI IgG in diluted Russell Viper Venom Time (dRVVT) assays, carried out using a PL dilution of 1:8 or 1:64, resulted in a delay in the onset of polymerization by 30-40 and 60-70s, respectively. Fibrin polymerization was complete after 250s for both anti-beta2GPI IgG and normal IgG. The inhibitory effect of the anti-beta2GPI antibodies was not observed in the presence of excess PL, as expected for LA. Anti-beta2GPI IgG increased the plateau level of polymerization when dRVVT was performed in the presence of 1.5 nM recombinant tissue plasminogen activator, but did not impair the fibrinolytic process, which was almost complete after 250 min. The autoantibodies did not delay the onset of fibrin polymerization in tests carried out using recombinant tissue factor. On the contrary, the autoantibodies enhanced polymerization in prothrombin time assays, and accelerated it in tissue thromboplastin inhibition tests, with no effect on fibrinolysis. These data provide evidence that anti-beta2GPI LA may act as either anticoagulants or procoagulants in different in vitro coagulation tests.