Investigations on the Nature of Hemophilia A

1963 ◽  
Vol 09 (01) ◽  
pp. 030-052 ◽  
Author(s):  
Eberhard Mammen
Keyword(s):  
Factor Viii ◽  
Human Plasma ◽  
Barium Sulfate ◽  
Freeze Drying ◽  
Room Temperature ◽  
Hemophilia A ◽  
Normal Human Plasma ◽  
Normal Human ◽  
And Storage ◽  
Prothrombin Activation

SummaryIn this paper an inhibitor is described that is found in hemophilic plasma and serum different from any till now described inhibitor. The inhibitor only inhibits prothrombin activation in the “intrinsic clotting systems”. This inhibitor is probably not present in normal human plasma or serum. It is destroyed by ether and freeze drying, is labile to acid and storage at room temperature. It is stable upon dialysis and has not been adsorbed on barium sulfate, aluminum hydroxide or kaolin. It precipitates at 50% v/v saturation with alcohol. The nature of this inhibitor seems to be a protein or lipoprotein.Factor VIII was isolated from hemophilic plasma. The amount isolated was the same as from normal plasma and the activity properties were not different. Hemophiliacs have normal amounts of factor VIII.

Prothrombin Fragment 1.2.3, A Major Product Of Prothrombin Activation In Human Plasma

1981 ◽  
Author(s):  
M J Rabiet ◽  
B Furie ◽  
B C Furie
Keyword(s):  
Sequence Analysis ◽  
Human Plasma ◽  
Gel Electrophoresis ◽  
Factor Xa ◽  
Normal Human Plasma ◽  
Terminal Sequence ◽  
Amino Terminal ◽  
Normal Human ◽  
Amino Terminal Sequence ◽  
Prothrombin Activation

The conversion of human prothrombin to thrombin is associated with a number of cleavage intermediates and products whose appearance and concentration are dependent upon the prothrombin activation conditions used. In the current investigation, the fragments of prothrombin which appear in normal human plasma after activation of the blood coagulation cascade were studied. Radioiodinated human prothrombin was added to platelet-poor relipidated normal human plasma and clotting initiated with Ca(II) and kaolin. The radiolabeled prothrombin cleavage products which formed were analyzed by polyacrylamide gel electrophoresis in the presence of dodecyl sulfate (SDS) and 2-mercaptoethanol (2-ME), A new product of prothrombin activation was observed. Its migration was more rapid than prethrombin 1 and slower than fragment 1.2. No previously identified products of prothrombin activation migrated to the same position in the gel.The previously unrecognized fragment was identified as fragment 1.2.3 as follows. Prothrombin was activated by factor Xa in the presence of Ca(II) and phospholipid. The desired product was isolated by absorption to and elution from barium citrate and by DEAE cellulose chromatography. This purified material, migrating identically with the unknown plasma product was homogeneous upon SDS gel electrophoresis with 2-ME. The amino terminal sequence of the isolated material was identical to that of prothrombin. Digestion of this material with either factor Xa or thrombin yielded as major products fragment 1.2 and fragment 1. (Fragment 2 and fragment 3 eluted from the gels under the conditions employed). Amino terminal sequence analysis of the factor Xa digestion products of the isolated material indicated three amino acid residues at each cycle. The sequences of fragment 1, fragment 2, and fragment 3 are consistent with this sequence analysis. On this basis we suggest that fragment 1.2.3 is a prominent product of prothrombin conversion to thrombin in plasma.

Human Protein C, Its Inhibitor, And The Combined Deficiency Of Factors V And VIII

1981 ◽  
Author(s):  
Richard A Marlar ◽  
John H Griffin
Keyword(s):  
Factor Viii ◽  
Human Plasma ◽  
Partial Thromboplastin Time ◽  
Protein C ◽  
Human Protein ◽  
Clotting Factor ◽  
Factor V ◽  
Normal Human Plasma ◽  
Normal Plasma ◽  
Normal Human

Human protein C (PC) is a vitamin K-dependent serine protease zymogen present in normal plasma. PC was activated with soluble thrombin, thrombin-Sepharose, trypsin-Sepharose or Russell’s viper venom. When activated protein C (APC) at 10 μg/ml was added to normal plasma for 3 min prior to assays, clotting was greatly inhibited. In prothrombin time, activated partial thromboplastin time, and partial thromboplastin time assays, the clotting times were prolonged from 24 to 68 sec, 57 to 616 sec, and 247 to > 1200 sec, respectively. Both calcium and phospholipid were needed for this potent anticoagulant activity. The thrombin time was unaffected. To test which coagulation factors in plasma were inactivated by APC, APC was added to normal human plasma for 3 min and residual clotting factor activities were assayed using appropriate deficient substrate plasmas. Only Factor V and Factor VIII activities were decreased by APC. PC and DFP-inhibited APC were not anticoagulant and did not decrease Factors V and VIII. APC hydrolyzed synthetic peptide substrates. The amidolytic activity of APC was inhibited by normal human plasma (50% of APC activity was lost in 10 min). APC inhibitory activity was not detectable in plasmas from 4 unrelated patients with combined Factor V/VIII deficiency whereas it was present in normal amounts in plasmas from patients with simple Factor V deficiency or Factor VIII deficiency. Thus, it appears that APC may regulate blood coagulation by enzymatically inactivating two critical cofactors, Factor Va and Villa. Moreover, APC itself may be regulated by a previously undescribed plasma protein inhibitor. It is suggested that the molecular basis for combined Factor V/VIII deficiency disease that exhibits simple autosomal recessive inheritance is a deficiency of APC inhibitor.

scholarly journals The Apparent Separation of Factor VIII Related Antigen and Coagulant Activity from von Willebrand Factor Activity by an Immunoadsorbent

1977 ◽  
Author(s):  
H. A. Cooper ◽  
D. Lee ◽  
M. A. Lamb ◽  
R. H. Wagner
Keyword(s):  
Factor Viii ◽  
Human Plasma ◽  
Human Factor ◽  
Solid Phase ◽  
Residual Activity ◽  
Normal Human Plasma ◽  
Human Factor Viii ◽  
Coagulant Activity ◽  
Plasma Fractions ◽  
Normal Human

An antibody was raised in rabbits to the small active fragment of human factor VIII. The antigen was obtained by Ca2+ dissociation of a human factor VIII preparation made from a multidonor pool of plasma. After two adsorptions with 0.1 volume of normal human plasma, the antibody neutralized the F. VIII coagulant activity of normal human plasma, but did not precipitate with any plasma or plasma fractions nor did it neutralize vWF activity as measured by ristocetin aggregation of fixed washed platelets. A solid phase immunochemical reagent was prepared by CNBr binding of the partially purified rabbit antibody to 1% agarose beads. Non-immune beads were similarly prepared with IgG fractions from a normal non-immunized rabbit. Using a batch technique the beads were studied for their ability to remove F. VIII coagulant, F. VIII Ag, and vWF activity from normal human plasma. Assay of the supernatant plasma after 2 hrs, 22°, from 10 replicate experiments gave the following results for residual activity, as per cent of non-immune bead control:F. VIII (37.5±4), F. VIII-Ag (30.8±9.7), and vWF (72.1±16). The experiment was repeated with 6 replicate samples with higher ratio of beads to plasma with essentially similar results. This unexpected separation of F. VIII-Ag from vWF activity prompted further investigation into how these activities are related to the molecular structure of F. VIII and vWF.

Performances of an Artificial Reagent for the One-stage Factor IX Assay

1975 ◽  
Vol 33 (03) ◽  
pp. 547-552 ◽  
Author(s):  
L Meunier ◽  
J. P Allain ◽  
D Frommel
Keyword(s):  
Human Plasma ◽  
Factor Ix ◽  
Severe Haemophilia ◽  
Normal Human Plasma ◽  
Haemophilia B ◽  
One Stage ◽  
Normal Human ◽  
The One

SummaryA mixture of adsorbed normal human plasma and chicken plasma was prepared as reagent for factor IX measurement using a one-stage method. The substrate was found to be specific for factor IX. Its performances tested on samples displaying factor IX activity ranging from <l%–2,500% compared favorably with those obtained when using the plasma of severe haemophilia B patients as substrate.

Plasma Components which Interfere with Ristocetin-induced Platelet Aggregation

1975 ◽  
Vol 33 (03) ◽  
pp. 540-546 ◽  
Author(s):  
Robert F Baugh ◽  
James E Brown ◽  
Cecil Hougie
Keyword(s):  
Platelet Aggregation ◽  
Human Plasma ◽  
Plasma Proteins ◽  
Binding Protein ◽  
Plasma Heating ◽  
Protein S ◽  
Fold Increase ◽  
Normal Human Plasma ◽  
Preliminary Examination ◽  
Normal Human

SummaryNormal human plasma contains a component or components which interfere with ristocetin-induced platelet aggregation. Preliminary examination suggests a protein (or proteins) which binds ristocetin and competes more effectively for ristocetin than do the proteins involved in ristocetin-induced platelet aggregation. The presence of this protein in normal human plasma also prevents ristocetin-induced precipitation of plasma proteins at levels of ristocetin necessary to produce platelet aggregation (0.5–2.0 mg/ml). Serum contains an apparent two-fold increase of this component when compared with plasma. Heating serum at 56° for one hour results in an additional 2 to 4 fold increase. The presence of a ristocetin-binding protein in normal human plasma requires that this protein be saturated with ristocetin before ristocetin-induced platelet aggregation will occur. Variations in the ristocetin-binding protein(s) will cause apparent discrepancies in ristocetin-induced platelet aggregation in normal human plasmas.

Analysis of Intrinsic Fibrinolysis in Human Plasma Induced by Dextran Sulfate

1992 ◽  
Vol 67 (04) ◽  
pp. 440-444 ◽  
Author(s):  
Hiroko Tsuda ◽  
Toshiyuki Miyata ◽  
Sadaaki Iwanaga ◽  
Tetsuro Yamamoto
Keyword(s):  
Molecular Weight ◽  
Human Plasma ◽  
Dextran Sulfate ◽  
Tissue Type ◽  
Factor Xii ◽  
Normal Human Plasma ◽  
High Molecular Weight Kininogen ◽  
Single Chain ◽  
Major Pathway ◽  
Normal Human

SummaryThe analysis of normal human plasma by fibrin autography revealed four species of plasminogen activator (PA) activity related to tissue-type PA, factor XII, prekallikrein and urokinase-type PA (u-PA). The u-PA activity increased significantly by incubating plasma with dextran sulfate. This increase was coincident with both the cleavage of factor XII and the complex formation of activated factor XII with its plasma inhibitors, which were determined by immunoblotting procedure. The dextran sulfate-dependent activation of u-PA required both factor XII and prekallikrein, but did not require either plasminogen or factor XI. High molecular weight kininogen was required only at a low concentration of dextran sulfate. Thus the results indicate that the factor XII and prekallikrein-mediated activation of single chain u-PA (scu-PA) operates as a major pathway of scu-PA activation in whole plasma in contact with dextran sulfate.

Presence and Possible Origin of ɛ(γ-Glutamyl)Lysine Isodipeptide in Human Plasma

1992 ◽  
Vol 67 (01) ◽  
pp. 060-062 ◽  
Author(s):  
J Harsfalvi ◽  
E Tarcsa ◽  
M Udvardy ◽  
G Zajka ◽  
T Szarvas ◽  
...  
Keyword(s):  
Human Plasma ◽  
Fibrinolytic Therapy ◽  
Normal Human Plasma ◽  
Normal Human ◽  
Hplc Technique ◽  
Significant Change

Summaryɛ(γ-glutamyl)lysine isodipeptide has been detected in normal human plasma by a sensitive HPLC technique in a concentration of 1.9-3.6 μmol/1. Incubation of in vitro clotted plasma at 37° C for 12 h resulted in an increased amount of isodipeptide, and there was no further significant change when streptokinase was also present. Increased in vivo isodipeptide concentrations were also observed in hypercoagulable states and during fibrinolytic therapy.

The Immunological Properties of Factor VIII

1966 ◽  
Vol 16 (03/04) ◽  
pp. 559-573 ◽  
Author(s):  
L Uszyński
Keyword(s):  
Factor Viii ◽  
Inhibitory Activity ◽  
Hemophilia A ◽  
Severe Form ◽  
Molecular Defect ◽  
Bovine Plasma ◽  
Human Factor Viii ◽  
Antigenic Properties ◽  
Coagulation Tests ◽  
Normal Human

SummaryRabbits immunized against human AHG fibrinogen-free preparations, were shown to produce anti-AHG antibodies. The inhibitory activity of these antibodies was tested by thromboplastin generation test, thrombelastography, and the specific anti-AHG antibodies neutralization test. The latter test permitted quantitative determination of antigenic form of factor VIII. The inhibitory activity of anti-FI-O-Ta serum resulted exclusively from the anti-AHG antibodies which in coagulation tests behaved like circulating anticoagulants directed against factor VIII.The anti-AHG antibodies were neutralizable by normal human serum or plasma even contained only trace of AHG activity after storage. There was no antigenic form of factor VIII in the severely affected patients with hemophilia A, von Willebrand’s disease nor in the normal plasma adsorbed on bentonite. The presented results suggest a molecular defect of factor VIII in patients with hemophilia A. The severe form of this disease depends, probably, on a major impairment of AHG biosynthesis, leading to changes in the antigenic properties of the molecule. The AHG from rabbit, porcine and bovine plasma respectively did not neutralize the anti-AHG antibodies formed in rabbits immunized against human factor VIII preparations.

A Chromogenic Factor X Assay With New Standardized Reagents

1979 ◽  
Author(s):  
P Friberger ◽  
C Lenne
Keyword(s):  
Human Plasma ◽  
Suitable Method ◽  
Chromogenic Substrate ◽  
Factor X ◽  
Normal Human Plasma ◽  
Normal Plasma ◽  
Normal Human

A recently published method for Factor X (FX) assay (1) utilizing Russel's Viper Venom (RVV) and a chromogenic substrate has been further investigated by testing a large number of parameters. This method has been considered as a suitable method for monitoring coumarol treatment (Bergström et al).The conditions for the activation of FX by purified preparations of the RVV have been studied as well as the conditions for FXa determination with a new chromogenic substrate Bz-Ile-Glu(γ-piperidyl)-Gly-Arg-pNA (S-2337). Both purified factors and normal plasma have been used. The effect of plasma inhibitors as well as the selectivity of the method has been studied.The reproducibility and stability of the different reagents and standards have been studied and found to be good.The amount of FXa activity obtained from normal human plasma has been titrated with FXa inhibitors of known purity.1) Aurell L. et al, Thromb. Res., 11, 595 (1977)2) Bergström et al, Thromb. Res., 12, 531 (1978)

Sign in / Sign up

Export Citation Format

Share Document