The Stability of Factor VIII in Heparinized Plasma

Summary

We studied the stability for 24 hr of factor VIII: C and factor Vm C:Ag in plasma collected in citrate, EDTA or heparin, and confirmed the previously reported two-phase decay of factor VIII:C in plasma when calcium ions have been chelated. We observed that plasma factor VIII:C is remarkably stable at 22° C (±2°) when normal calcium ion concentrations are maintained. The loss of activity of factor VIII:C between one and 24 hr after blood collection was on average only 0.5% per hr for heparinized plasma. There was also an apparent loss of factor VIII C:Ag in plasma where calcium ions had been removed, compared with factor VIII C:Ag in heparinized plasma. However, a comparison of one-site and two-site assays suggested that calcium chelating agents may lead to factor VIII C:Ag levels being under-estimated when one-site fluid-phase assays are employed.

To confirm the action of calcium ions in maintaining factor VIII:C stability, we carried out a series of experiments where calcium chloride was added four hr after blood collection to plasma anticoagulated by a mixture of citrate plus heparin; we observed total recovery of factor VIII:C activity within four hr.

The stability of factor VIII:C, even at room temperature, in the presence of physiological calcium ion concentrations has implications for manufacturers of factor VIH concentrates and cryo-precipitates.

Keywords

Factor VIII stability - Heparin - Plasma - Calcium

• References

• 1 Stibbe J, Hemker HC, Creveld SV. The inactivation of factor VIII in vitro. Thrombos Diathes Haemorrh 1972; 27: 43-58
• 2 Rock GA, Cruickshank WH, Tackaberry ES, Palmer DS. Improved yields of factor Vm from heparinised plasma. Vox Sang 1979; 36: 294-300
• 3 Rock GA, Cruickshank WH, Tackaberry ES, Ganz PR, Palmer DS. Stability of VIILC in plasma: the dependence on protease activity and calcium. Thromb Res 1983; 29: 521-535
• 4 Biggs R. Human Blood Coagulation, Haemostasis and Thrombosis, 2nd edit. Blackwell, Oxford: 1976. p 681
• 5 Nilsson IM, Kirkwood TB L, Barrowcliffe TW. In vivo recovery of factor VIII: A comparison of one-stage and two-stage assay methods. Thromb Haemostas 1979; 42: 1230-1239
• 6 Lazarchick J, Hoyer LW. Immunoradiometric measurement of the factor VIII procoagulant antigen. J Clin Invest 1978; 62: 1048-1052
• 7 Fine JM, Steinbuch M. A simple technique for the isolation of monoclonal IgG and IgA. Rev Eur Etud Clin Biol 1970; 15: 1115-1121
• 8 Reisner HM, Barrow ES, Graham JB. Radioimmunoassay for coagulant factor VUI-related antigen (F Vm C:Ag). Thromb Res 1979; 14: 235-239
• 9 Hunter WM, Greenwood FC. Preparation of iodine-131 labelled human growth hormone of high specific activity. Nature 1962; 294: 495-496
• 10 Peake IR, Bloom AL. Immunoradiometric assay of procoagulant factor VIII antigen in plasma and serum and its reduction in haemophilia. Lancet 1978; 01: 473-475
• 11 Rotblat F, Tuddenham EG D. Immunologic studies of factor Vin coagulant activity (VIII :C) I. Assay based on a haemophilic and an acquired antibody to factor VIILC. Thromb Res 1981; 21: 431-435
• 12 Rock GA, Tackaberry ES, Palmer DS. Factors affecting the relative distribution of high and low molecular weight forms of factor Vin. Thromb Res 1979; 14: 573-587
• 13 Rock GA, Palmer DS, Tackaberry ES, Cruickshank WH. The presence of high and low molecular weight forms of factor VIII in heparinised plasma. Thromb Res 1978; 13: 85-96
• 14 Seghatchian MJ, Nilsson IM, Holmberg L, Miller-Andersson M. Molecular size distribution of factor VIII in native plasma. Thromb Res 1979; 14: 589-598
• 15 Damus PS, Hicks M, Rosenberg RD. Anticoagulant action of heparin. Nature 1973; 246: 355-357
• 16 Kakkar VV. Prevention of venous thromboembolism. In: Haemostasis and Thrombosis. Bloom AL, Thomas DP. (eds). Churchill Livingstone: 1981. pp 669-683
• 17 Thomas DP, Sagar S, Stamatakis JD, Maffei FH A, Erdi A, Kakkar VV. Plasma heparin levels after administration of calcium and sodium salts of heparin. Thromb Res 1976; 09: 241-248