Fibrinogen/Fibrin Degradation Products in Defibrination Syndrome Analysed on the Basis of Molecular Weight on Gel Exclusion Chromatography

Exclusion Chromatography ◽

D Antigen ◽

Stabilized Fibrin

1) Fresh citrated plasmas of cases of defibrination syndrome were chromatographed on Biogel A-5 m column, which revealed a two-peaked elution profile of fibrinogen-related antigen with reproducibility. The first peak was eluted at the Vo of the column and the second one at Ve of the early FDP. It was found in the preliminary experiments in vitro, that the big molecular FDP was formed exclusively in the process of fibrin clot lysis, irrespective if it was non-stabilized or stabilized.2) FDP in thesen patients’ serums, after thorough digestion with plasmin, were chromatographed on Sephadex G200 column. A reproducible elution profile of FDP-D antigen revealed single peak, of which Ve was corresponded with that of FDP-D monomer derived from fibrinogen or non-stabilized fibrin.It was postulated that the FDP in plasmas of defibrination syndrome were mainly originated from the fibrinogen or non-stabilized fibrin formed in the process of the diseases.

Identification of D-Dimer-E Complex in Disseminated Intravascular Coagulation (DIC)

10.1055/s-0039-1684840 ◽

1979 ◽

Author(s):

A.N. Whitaker ◽

E.A. Rowe ◽

P.P. Masci ◽

P.J. Gaffney

Keyword(s):

Gel Electrophoresis ◽

Degradation Products ◽

Polyacrylamide Gel Electrophoresis ◽

Stable Complex ◽

Pneumococcal Sepsis ◽

D Antigen ◽

D Dimer

D-dimer (D2), a product of the plasmin lysis of cross-linked (XL) fibrin, but not of non-XL fibrin or fibrinogen, has been identified in the plasma of patients with DIC due to amniotic fluid embolism. In vitro, D is involved with fragment E as a stable complex (D2-E) but D2 -E has not been identified in vivo before. Fibrin degradation products (FDP) were studied in a patient having fulminant postsplenectomy pneumococcal sepsis and DIC, by immunoprecipitation with anti-fibrinogen (f) and anti-fragment E and characterization by SDS polyacrylamide gel electrophoresis (PAGE). With both antisera soluble HMW fibrin complexes, D2 and E but no X, Y or D were obtained from serum. D2 and E were identified in the supernatant after removing partially XL HMW complexes and fibrinogen from plasma with 2.5 M β-alanine. The presence D antigen in the D2-E complex precipitated by monospecific anti-E was confirmed by crossed Ag-Ab electrophoresis. Crossed Ag-Ab electrophoresis of serum in agarose gave E peaks of slow mobility and no fast-moving free E was found. Thus, D2-E complex exists in vivo and its easy identification, proving the lysis of XL fibrin, would be of value in studying thrombosis. D2-E complex has been identified in other patients with sepsis but at lower concentrations than described above.

The Fibrin Assay Comparison Trial (FACT)

10.1055/s-0037-1615652 ◽

2001 ◽

Vol 85 (04) ◽

pp. 671-678 ◽

Cited By ~ 82

Author(s):

Sybille Zips ◽

Hanimsah Ergül ◽

Dieter Heene ◽

Carl-Erik Dempfle ◽

Keyword(s):

Molecular Weight ◽

High Molecular Weight ◽

Degradation Products ◽

Plasma Samples ◽

Comparison Trial ◽

Clinical Conditions ◽

D Dimer

SummaryAlthough D-dimer has gained widespread clinical use as a parameter for detection of in vivo fibrin formation, the issue of standardization of D-dimer assays remains to be resolved. The FACT study was performed to generate basic data for development of calibrators and standard preparations.A set of 86 samples, including plasma samples from patients with DIC, DVT, and other clinical conditions, serial dilutions of pooled plasma samples, and plasma samples containing fibrinogen- and fibrin derivatives, were distributed to 12 manufacturers of D-dimer assays.D-dimer assays differ concerning specificity for crosslinked fibrin, and preference for either high molecular weight fibrin complexes, or low molecular weight fibrin degradation products. Terminal plasmin digests of fibrin clots for calibration produce aberrant results in some assays, especially those with preference for high molecular weight crosslinked fibrin derivatives. The best conformity is achieved by the use of pooled plasma samples from patients with high levels of D-dimer antigen in plasma. In vitro preparations containing a comparable composition of fibrin derivatives to clinical plasma samples may also serve as reference material.

The cofactor role of protein S in the acceleration of whole blood clot lysis by activated protein C in vitro

Blood ◽

10.1182/blood.v67.4.1189.1189 ◽

1986 ◽

Vol 67 (4) ◽

pp. 1189-1192 ◽

Cited By ~ 60

Author(s):

NJ de Fouw ◽

F Haverkate ◽

RM Bertina ◽

J Koopman ◽

A van Wijngaarden ◽

...

Keyword(s):

Whole Blood ◽

Protein C ◽

Degradation Products ◽

Blood Clot ◽

Activated Protein C ◽

Protein S ◽

Tissue Type ◽

Clot Lysis ◽

Fibrin Degradation Products

Abstract The effect of purified human activated protein C (APC) and protein S on fibrinolysis was studied by using an in vitro blood clot lysis technique. Blood clots were formed from citrated blood (supplemented with 125I-fibrinogen) by adding thrombin and Ca2+-ions; lysis of the clots was achieved by adding tissue-type plasminogen activator. The release of labeled fibrin degradation products from the clots into the supernatant was followed in time. We clearly demonstrated that APC accelerates whole blood clot lysis in vitro. The effect of APC was completely quenched by antiprotein C IgG, pretreatment of APC with diisopropylfluorophosphate, and preincubation of the blood with antiprotein S IgG. This demonstrates that both the active site of APC and the presence of the cofactor, protein S, are essential for the expression of the profibrinolytic properties. At present, the substrate of APC involved in the regulation of fibrinolysis is not yet known. Analysis of the radiolabeled fibrin degradation products demonstrated that APC had no effect on the fibrin cross-linking capacity of factor XIII.

Alteration of fibrin network by activated protein C

Blood ◽

10.1182/blood.v83.9.2541.2541 ◽

1994 ◽

Vol 83 (9) ◽

pp. 2541-2548 ◽

Cited By ~ 18

Author(s):

A Gruber ◽

E Mori ◽

GJ del Zoppo ◽

L Waxman ◽

JH Griffin

Keyword(s):

Protein C ◽

Degradation Products ◽

Activated Protein C ◽

Relative Number ◽

Relative Mass ◽

Clot Lysis ◽

Rate Of Increase ◽

Fibrin Network

Abstract The antithrombotic plasma enzyme, activated protein C (APC), may play a role in thrombolysis. In vitro, acceleration of clot lysis by APC depends on its ability to inhibit the activation of prothrombin. The effect of APC on the assembly and dispersion of fibrin network was studied using turbidimetry, plasmin digestion of fibrin, and electron microscopy of plasma clots. The addition of APC before clotting but not after clotting accelerated clot lysis. The rate of increase in the turbidity of clotting plasma was reduced by APC. The turbidity of plasma clots containing APC was directly related to the clot lysis time. Fibrin from plasma clots that were formed in the presence of APC yielded less fibrin degradation products than fibrin from clots without added APC. Furthermore, APC reduced the diameter and relative number of fibrin fibers in plasma clots during gel assembly. We propose that APC may enhance the efficacy of thrombolysis by reducing the relative mass of fibrin within maturing thrombi.

In Vitro Formation of Fibrin-Monomer Complexes in the Presence of Isotope Labelled Fibrinogen-Fibrin Degradation Products. An Agarose Gel Filtration Study

10.1055/s-0039-1689556 ◽

1975 ◽

Author(s):

F. Asbeck ◽

van de J. Loo

Keyword(s):

Molecular Weight ◽

Complex Formation ◽

Degradation Products ◽

Gel Filtration ◽

Agarose Gel ◽

Fibrinogen Degradation Products ◽

Molecular Weights ◽

Fibrin Monomer

Human citrated plasmas were mixed with purified 131I-fibrinogen and 131I-fibrinogen degradation products (FDP) or 125I-fibrin degradation products (fdp). After incubation with small amounts of thrombin (0.01–0.02 imits/ml Pl.), these mixtures were gel filtrated on Biogel A5m columns and the elution patterns of the 131I- and -labelled materials were determined.In control experiments without thrombin incubation, no complex formation between fibrinogen, FDP or fdp in citrated plasmas could be detected. This was even true for fdp with a higher molecular weight than fibrinogen.After thrombin incubation, up to 11% fibrin-monomer complexes were formed. Irrespective of their molecular weights, labelled fdp were not incorporated into these complexes.Only large FDP – presumably derivative X – did partially copolymerize with fibrin-monomer complexes in citrated plasmas.

The cofactor role of protein S in the acceleration of whole blood clot lysis by activated protein C in vitro

Blood ◽

10.1182/blood.v67.4.1189.bloodjournal6741189 ◽

1986 ◽

Vol 67 (4) ◽

pp. 1189-1192

Author(s):

NJ de Fouw ◽

F Haverkate ◽

RM Bertina ◽

J Koopman ◽

A van Wijngaarden ◽

...

Keyword(s):

Whole Blood ◽

Protein C ◽

Degradation Products ◽

Blood Clot ◽

Activated Protein C ◽

Protein S ◽

Tissue Type ◽

Clot Lysis ◽

Fibrin Degradation Products

The effect of purified human activated protein C (APC) and protein S on fibrinolysis was studied by using an in vitro blood clot lysis technique. Blood clots were formed from citrated blood (supplemented with 125I-fibrinogen) by adding thrombin and Ca2+-ions; lysis of the clots was achieved by adding tissue-type plasminogen activator. The release of labeled fibrin degradation products from the clots into the supernatant was followed in time. We clearly demonstrated that APC accelerates whole blood clot lysis in vitro. The effect of APC was completely quenched by antiprotein C IgG, pretreatment of APC with diisopropylfluorophosphate, and preincubation of the blood with antiprotein S IgG. This demonstrates that both the active site of APC and the presence of the cofactor, protein S, are essential for the expression of the profibrinolytic properties. At present, the substrate of APC involved in the regulation of fibrinolysis is not yet known. Analysis of the radiolabeled fibrin degradation products demonstrated that APC had no effect on the fibrin cross-linking capacity of factor XIII.

Availability of the Bβ(15-21) Epitope on Cross-Linked Human Fibrin and Its Plasmic Degradation Products

10.1055/s-0038-1648443 ◽

1992 ◽

Vol 67 (03) ◽

pp. 335-340 ◽

Cited By ~ 7

Author(s):

Fabian Chen ◽

Edgar Haber ◽

Gray R Matsueda

Keyword(s):

Molecular Weight ◽

Calcium Ions ◽

Degradation Products ◽

Plasminogen Activators ◽

Fibrin Clot ◽

Clot Lysis ◽

Low Molecular Weight ◽

Potential Utility ◽

Antibody Preparations ◽

Fibrin Clots

SummaryThe binding of radiolabeled monoclonal antifibrin antibody 59D8 (specific for fibrin but not fibrinogen) to a series of degraded fibrin clots showed that the availability of the Bβ(15-21) epitope (against which 59D8 had been raised) was inversely proportional to the extent of clot lysis. Examination of digest supernatants revealed that the Bβ(15-21) epitope was released from clots as a high molecular weight degradation product in the presence of calcium ions but that the generation of low molecular weight peptides occurred in the absence of calcium ions. To address the question of epitope accessibility, we compared levels of fibrin clot binding among four radioactively labeled antibodies: antifibrin monoclonal antibody 59D8, two antifibrinogen monoclonal antibodies that cross-reacted with fibrin, and an affinity-purified polyclonal antifibrinogen antibody. We expected that the antifibrinogen antibodies would show enhanced binding to clots in comparison with the antifibrin antibody. However, the epitope accessibility experiments showed that all four antibody preparations bound fibrin clots at comparable levels. Taken together, these studies demonstrated that one fibrin-specific epitope, Bβ(15-21), remains available on clots as they undergo degradation by plasmin and, importantly, that the epitope is not solubilized at a rate faster than the rate at which the clot is itself solubilized. The availability of the Bβ(15-21) epitope during the course of plasminolysis assures the potential utility of antifibrin antibodies such as 59D8 for detecting thrombi and targeting plasminogen activators.

Thrombolysis with Streptokinase in Rabbits Dose Response, Fibrin-clot Specificity and Laboratory Evaluation of Fibrinolytic Effect

10.1055/s-0038-1649160 ◽

1974 ◽

Vol 31 (02) ◽

pp. 265-272

Author(s):

Andrzej Nowak ◽

Victor Gurewich

Keyword(s):

Animal Model ◽

Thrombolytic Therapy ◽

Dose Response ◽

Degradation Products ◽

Fibrin Clot ◽

Laboratory Evaluation ◽

Clot Lysis ◽

Excellent Correlation ◽

High Degree

SummaryAn animal model was used in which venous thromboemboli of reproducible size could be formed. The effects of SK 500–50,000 units per hour were evaluated and compared to those of saline. Proportionately the greatest clot lysis occurred with the lowest doses of SK. In contrast to thrombolysis, significant fibrinogenolysis did not occur in any of the animals indicating a high degree of specificity of SK-induced activator for rabbit fibrin. An excellent correlation between the concentration of fibrin degradation products (fdp) measured by the serial dilution protamine sulfate test and the extent of clot lysis was found suggesting this to be a useful method for monitoring the effects of thrombolytic therapy. A method for the laboratory distinction of fdp from fibrin monomer was demonstrated.

Alteration of fibrin network by activated protein C

Blood ◽

10.1182/blood.v83.9.2541.bloodjournal8392541 ◽

1994 ◽

Vol 83 (9) ◽

pp. 2541-2548

Author(s):

A Gruber ◽

E Mori ◽

GJ del Zoppo ◽

L Waxman ◽

JH Griffin

Keyword(s):

Protein C ◽

Degradation Products ◽

Activated Protein C ◽

Relative Number ◽

Relative Mass ◽

Clot Lysis ◽

Rate Of Increase ◽

Fibrin Network

The antithrombotic plasma enzyme, activated protein C (APC), may play a role in thrombolysis. In vitro, acceleration of clot lysis by APC depends on its ability to inhibit the activation of prothrombin. The effect of APC on the assembly and dispersion of fibrin network was studied using turbidimetry, plasmin digestion of fibrin, and electron microscopy of plasma clots. The addition of APC before clotting but not after clotting accelerated clot lysis. The rate of increase in the turbidity of clotting plasma was reduced by APC. The turbidity of plasma clots containing APC was directly related to the clot lysis time. Fibrin from plasma clots that were formed in the presence of APC yielded less fibrin degradation products than fibrin from clots without added APC. Furthermore, APC reduced the diameter and relative number of fibrin fibers in plasma clots during gel assembly. We propose that APC may enhance the efficacy of thrombolysis by reducing the relative mass of fibrin within maturing thrombi.

A Monoclonal Antibody-Based Enzyme Immunoassay for Fibrin Degradation Products in Plasma

10.1055/s-0038-1642777 ◽

1988 ◽

Vol 59 (02) ◽

pp. 310-315 ◽

Cited By ~ 64

Author(s):

P W Koppert ◽

E Hoegee-de Nobel ◽

W Nieuwenhuizen

Keyword(s):

Enzyme Immunoassay ◽

Degradation Products ◽

Blood Clot ◽

Tissue Type ◽

Type Plasminogen Activator ◽

Strong Positive Reaction ◽

Sandwich Type ◽

Capture Antibody

SummaryWe have developed a sandwich-type enzyme immunoassay (EIA) for the quantitation of fibrin degradation products (FbDP) in plasma with a time-to-result of only 45 minutes.* The assay is based on the combination of the specificities of two monoclonal antibodies (FDP-14 and DD-13), developed in our institute. FDP-14, the capture antibody, binds both fibrinogen degradation products (FbgDP) and FbDP, but does not react with the parent fibrin(ogen) molecules. It has its epitope in the E-domain of the fibrinogen molecule on the Bβ-chain between amino acids 54-118. Antibody DD-13 was raised using D-dimer as antigen and is used as a tagging antibody, conjugated with horse-radish peroxidase. A strong positive reaction is obtained with a whole blood clot lysate (lysis induced by tissue-type plasminogen activator) which is used as a standard. The EIA does virtually not detect FbgDP i. e. purified fragments X, Y, or FbgDP generated in vitro in plasma by streptokinase treatment. This indicates that the assay is specific for fibrin degradation products.We have successfully applied this assay to the plasma of patients with a variety of diseased states. In combination with the assay previously developed by us for FbgDP and for the total amount of FbgDP + FbDP (TDP) in plasma, we are now able to study the composition of TDP in patients plasma in terms of FbgDP and FbDP.