Studies on the Neutralization of Rabbit Antibodies to Human Factor VIII

SummaryThe capacity of normal and haemophilic cryoprecipitates to neutralize the anticoagulant effect of rabbit antibodies to human factor VIII (anti-VIII) was assessed using a quantitative estimation of antibody. About 4 times as much anti-VIII could be neutralized by normal factor VIII as was required to neutralize clotting activity. This suggests that there are probably several antigenic sites intimately associated with factor VIII clotting activity, quite apart from any antigenic sites which may be detected using antibodies directed against other components of the factor VIII complex. The neutralizing capacity of factor VIII was only slightly greater for the rabbit antibodies employed in this study than has been previously reported for antibodies of human origin, thus indicating only minor differences in specificities. Additional evidence in support of this concept was the finding that cryoprecipitates prepared from haemophilic plasmas previously recognized as either lacking or possessing the capacity to neutralize antibodies of human origin neutralized least or most quantities of rabbit antibodies, respectively.

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The Specificity of Antibodies to Factor VIII Produced in the Rabbit after Immunization with Human Cryoprecipitate

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648107 ◽

1973 ◽

Vol 29 (03) ◽

pp. 652-660 ◽

Cited By ~ 3

Author(s):

P. B. A Kernoff ◽

C. R Rizza

Keyword(s):

Factor Viii ◽

Human Factor ◽

Neutralizing Antibody ◽

Normal Subjects ◽

Factor Viii Activity ◽

Marked Rise ◽

Antigenic Sites ◽

Level Of Activity ◽

Human Factor Viii ◽

The Relationship

SummaryThe relationship between the activity-neutralizing and precipitating activities of rabbit antibody to human factor VIII was studied by measuring the changes in levels of the two activities in two sensitized rabbits after stimulation with cryoprecipitates prepared from the plasmas of normal subjects, haemophiliacs and patients with von Wil- lebrand’s disease. Injection of cryoprecipitates prepared from plasmas with a detectable level of factor VIII clotting activity was followed by a marked rise in the level of activity-neutralizing antibody, whereas there was no rise or a continued fall in level after injection of cryoprecipitates prepared from haemophilic plasmas without detectable factor VIII activity. The level of precipitating antibody rose markedly after injection of normal and haemophilic cryoprecipitates, but little or not at all after cryoprecipitates prepared from the plasma of the patients with von Willebrand’s disease. It is suggested that the specific antigenic sites associated with factor VIII clotting activity were not present in cryoprecipitates prepared from haemophilic plasmas without detectable factor VIII activity, and also that antibodies of two different specificities could be detected.

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The Fractionation Properties of Human Factor VIII (Antihaemophilic Factor)

Thrombosis and Haemostasis ◽

10.1055/s-0038-1654503 ◽

1960 ◽

Vol 04 (02) ◽

pp. 253-260 ◽

Cited By ~ 1

Author(s):

Franco Gobbi

Keyword(s):

Factor Viii ◽

Human Factor ◽

Maximum Yield ◽

Factor Vii ◽

Precipitation Method ◽

Salting Out ◽

Human Factor Viii ◽

Plasma Factor ◽

Two Factors ◽

Thromboplastic Activity

SummaryThe fractionation properties of human Factor VIII (antihaemophilic factor, AHF, antihaemophilic globulin) have been studied using a plasma of congenital afibrinogenaemia as a starting material.From a fibrinogen-free plasma, Factor VIII does not precipitate with ethanol at a final concentration of 8%; on the contrary the maximum yield is reached at an ethanol concentration of 25%.With a precipitation method carried out by a one to ten dilution of plasma with distilled water and acidification by N/10 hydrochloric acid to a pFI 5.2, Factor VIII does not precipitate with the euglobulin fraction; when normal plasma is used, such a precipitation is almost complete.With the salting-out fractionation method by ammonium sulphate, Factor VIII precipitates at a concentration between 25 and 33% of saturation either from fibrinogen-free and from normal human plasma.A non-specific thromboplastic activity appears in the fractions prepared by every method. This activity, which is probably due to the activation of seric accelerators, is easily removed by Al(OH)s adsorption. Thus, in order to insure the specificity of Factor VIII assays, the preliminary adsorption of the fractions is indispensable before testing their antihaemophilic activity.Fibrinogen and Factor VIII have different and definite precipitation patterns. When these two factors are associated the fractionation properties of AHF appear quite modified, showing a close similarity to those of fibrinogen. This fact can explain the technical difficulties encountered in the attempt to purify the antihaemophilic factor, and the lack of reproducible procedures for removing fibrinogen without affecting Factor VII.

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Impaired Prothrombin Consumption in Bernard-Soulier Syndrome Is Corrected In Vitro by Human Factor VIII

Thrombosis and Haemostasis ◽

10.1055/s-0038-1655972 ◽

1997 ◽

Vol 77 (02) ◽

pp. 383-386 ◽

Cited By ~ 8

Author(s):

S Bellucci ◽

J P Girma ◽

M Lozano ◽

D Meyer ◽

J P Caen

Keyword(s):

Factor Viii ◽

Human Factor ◽

Platelet Membrane ◽

Giant Platelets ◽

Prolonged Bleeding Time ◽

Human Factor Viii ◽

Von Willebrand ◽

Willebrand Factor ◽

Bernard Soulier Syndrome

SummaryThe Bernard-Soulier syndrome (BSS) is characterized by thrombocytopenia with giant platelets, a prolonged bleeding time with defective platelet adhesion to the subendothelium related to a defect in platelet membrane glycoprotein lb (GPIb) and a decreased prothrombin consumption. The mechanism of the latter abnormality remains unknown. In this study, we showed that this defect was corrected by the addition of purified human factor VIII (FVIII) to blood from four patients with BSS. The correction of prothrombin consumption was almost complete at concentrations between 1.5 and 3 IU/ml of FVIII procoagulant activity (VIII.'C) and partially abolished by a monoclonal antibody which neutralizes VIII:C. This correction was specific for FVIII and was not observed after addition of purified human FIX. It was obtained, in the same magnitude range, with FVIII complexed to von Willebrand factor (vWF) but not with free vWF. These data provide a new insight into the knowledge of the physiological interaction between the platelet membrane and the vWF-FVIII complex facilitating plasma coagulation activation and may lead to helpful therapeutic advances.

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Synthesis, processing, and secretion of recombinant human factor VIII expressed in mammalian cells.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(18)68793-9 ◽

1988 ◽

Vol 263 (13) ◽

pp. 6352-6362 ◽

Cited By ~ 9

Author(s):

R J Kaufman ◽

L C Wasley ◽

A J Dorner

Keyword(s):

Factor Viii ◽

Mammalian Cells ◽

Human Factor ◽

Human Factor Viii

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Immunopurification of Human Factor VIII/vWF Complex from Plasma

Thrombosis and Haemostasis ◽

10.1055/s-0038-1647497 ◽

1988 ◽

Vol 59 (03) ◽

pp. 364-371 ◽

Cited By ~ 10

Author(s):

Odile Mejan ◽

Vincent Fert ◽

Maryléne Delezay ◽

Michel Delaage ◽

Rose Cheballah ◽

...

Keyword(s):

Factor Viii ◽

Human Factor ◽

Specific Activity ◽

Coupling Method ◽

Purification Process ◽

Solid Supports ◽

Coupling Methods ◽

Human Factor Viii ◽

Alkaline Conditions

SummaryIn this study we describe a process for immunopurification of FVIII/vWF complex directly from plasma. A mAb against vWF has been selected that is able to bind, under physiologic conditions, the FVIII/vWF complex and to release it in slightly alkaline conditions while preserving its activity.After investigating the influence of solid supports and of coupling methods on the recovery of active FVIII we produced an immunoadsorbent by immobilisation of the selected mAb onto a Sephacryl S-1000 support using a benzoquinone coupling method. With this immunoadsorbent we developed a purification process directly from plasma with an excellent recovery (50%) of both FVIII and vWF activities. The product obtained is very enriched (the FVIII: C specific activity is 20 IU/mg of protein) and is stable after lyophilization.

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