Gel Structure And Clotting Time

Abstract

Ferry and Morrisson found on the basis of the mechanical and optical properties of fibrin gels that they ranged between two extreme types: fine or transparent and coarse or opaque. We have now developed a device permitting studies of flow properties of gels. The gels are formed in standardized cups (1.47×2.5 cm) which are fitted into a special holder attached to a reservoir for application of buffer solution for the flow measurement. Gels were formed by mixing thrombin or Batroxobin with fibrinogen solutions of different protein concentrations (1-4 g/1), pH (6.5-8.2) and ionic strengths (0.1-0.23). All these studies were in addition performed at different enzyme concentrations. The gels were left in the cups for complete gelation to occur (1-2 hours) and subsequently the same buffer as used for gel formation was percolated through the gels at room temperature and a constant pressure gradient. In parallell experiments the formation and optical properties of the gels formed was studied by turbidity measurement at 450 nm. After a lagtime rapid increase in turbidity occurred. The lag-time was defined as the “clotting-time”.We found that for both enzymes the flow is related to the optical properties of the gels, being fast for the opaque and slow for transparent gels. Fast flow is thus favoured by low pH and low ionic strength. The flow rate is furthermore inversely related to the gel concentration. Of particular importance was the finding that under all conditions so far studied there was, within a wide range, a direct relationship between clotting time and flow rate. This suggests that the final structure of the gels is determined by events preceeding the gel formation. These events are enzymatic activation and alignement of activated molecules. Gel formation is therefore very likely an ordered process resembling that of crystallization. Batroxobin and thrombin gels having the same clotting times displayed different flow properties suggesting that different types of polymers are formed on release of one (FPA) and two (FPA + FPB) activation peptides, respectively.