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33. FIBRINOGEN DEGRADATION PRODUCTS AND A FIBRINOGEN ASSAY BASED ON CLOTTING KINETICS+

M.B. Donati”, J. Vermylen and M. Verstraete

Laboratory of Blood Coagulation (Head : hoJ Dr. M. Verstraete) Department of Internal Medicine (Head : Pro$ Dr. J. Vandenbroucke)

University of Leuven, Belgium

It has already been reported by many workers that some fragments resulting from progressive proteolysis of human fibrinogen by human plasmin can inhibit the fibrinogen-to-fibrin conversion by reducing the polymerization rate of fibrin monomers and modifying some physical properties of the clot (Bang et al. 1962, Hirsh et al. 1965).

Based on this observation, Hirsh et al. (1965) suggested that all fibrinogen assays based on clotting kinetics would give inaccurate results if the tested plasma contains fibrinogen proteolysis products.

This is also the reason why Fletcher et al. (1962) have introduced a modification of the procedure of Ratnoff and Menzie (1 95 l ) , which includes a 12-hour coagulation period, to measure the fibrinogen concentration in plas- mas containing fibrinogen degradation pro- ducts.

* On leave of absence from the Laboratory o f Blood Coagulation (Prof. B. Bizzi), Department of Inter- nal Medicine (Prof. R. Breda), Universiti Cattolica, Roma (Italy). Fellowship of the Minister0 della Pubblica Istru- zione, Roma.

f This investigation was supported by grant No 1216 of the Fonds voor Geneeskundig Wetenschappelijk Onderzoek. Brussel.

A fibrinogen assay based on clotting kinetics and derived from the assay procedure of Clauss (1957) has been developed in this laboratory several years ago (Vermylen et al. 1963a) ; the clotting time is measured in the presence of low fibrinogen levels (usually obtained by diluting the sample) and of very high concentrations of thrombin. In these conditions the measured clotting time has been found to be closely related to the concentration of fibrinogen. As the proteolysis step is reduced to a constant minimum, the test measures exclusively poly- merization upto the stage of visible fibrin formation. For this reason, the test is named Fibrin Polymerization Time-test (FPT-test).

As shown in figure 1, when this test is performed to measure the fibrinogen content of plasma from normal subjects, complete agree- ment between this rapid and simple method and the results obtained with Blomback’s method (1958) is found, which must be expected as the calibration of the FPT-test was performed using Blomback’s method (Vermy- len et al. 1963b). However, when the FPT-test is performed on plasma containing an increased level of fibrinogen degradation products, for instance during hyperplasminemia induced by streptokinase treatment (Verstraete e t al. 1971), a much lower fibrinogen content is found using the FPT-test than Blomback’s method (figure 1).

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Acknowledgements

We wish to thank Miss Arlette Verhaegen and Miss Trees Vancoetsem for their skilful technical assistance.

REFERENCES

Bang, N.U., Fletcher, A.P., Alkjaersig, N. and Sherry, S. (1 962) “Pathogenesis of the coagulation defect developing during pathological plasma proteolytic (“fibrinolytic”) states. I l l . Demonstration of abnor- mal clot. structure by electron microscopy”. J. Clin. Invest, 4 I . 9 35-948.

Blomback, B. ( I 958) “On the properties of fibrinogen and fibrin”. Arkiv Kemi 12, 99113.

Clauss, A. ( 1957) “Cerinnungsphysiologische Schnell- methode zur Bestimmung des Fibrinogens” Acto Haemat. 17, 237-246.

Fletcher, A.P., Alkjaersig, N. and Sherry, S. (1962) “Pathogenesis of the coagulation defect developing during pathological plasma proteolytic (“fibrino- lytic”) states. 1. The significance of fibrinogen proteolysis and circulating fibrinogen breakdown products”. J. Clin. Invest. 41, 896-916.

Hirsh, J., Fletcher, A.P. and Sherry S. (1965) “E.ffect of fibrin and fibrinogen proteolysis products on clot physical properties”. Amer. J. Physiol. 209,

Ratnoff, O.D. and Menzie, C. (1951) “A new method for the determination of fibrinogen in small samples of plasma”. J.Lab.Clin.Med. 37, 316-320.

Vermylen, C., De Vreker, R.A. and Verstraete, M. (1963a) “A quick quantitative enzymatic fibrinogen assay method : the fibrin polymerization time (FFT)”. Clin. Chim. Acto 8, 41 8-424.

Vermylen, C., De Vreker, R.A. and Verstraete, M. (1963b)“Usefulness of and a simple method for serial successive fibrinogen determinations in human pathology’.’Thrombos. Diathes haemorrhlo,

Verstraete, M., Vermylen, J. and Donati, M.B. (1971) “The effect of streptokinase infusion on chronic arterial occlusions and stenoses”. Ann.lnt.Med. 74,

4 15-424.

239-242.

3 17-3 8 2.

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0 100 Z O O 300 4 0 0 500 6 0 0 100 100 900 OLOWOICK T F S T mpX

Figure 1

Correlation between the results of fibrinogen determi- nation with the FPT-test and with Blomback’s method in normal plasma and plasma obtained from streptokinase-treated patients with increased FDP in serum.

These results show that the FPT-test is influenced both by the fibrinogen concen- tration and by the level of fibrinogen degradation products, when these are present. This may however not necessarily be a disadvantage, provided interpretation is done with caution. Indeed, the FPT-test may give a valuable indication of the amount of fibrinogen available for hemostasis in clinical situations with primary or secondary fibrin(0gen)olysis.