Bovine fibrinogen aggregates: electron microscopic observations of quasi-globular structures

Electron microscopic observations of bovine fibrinogen preparations were made following negative staining with uranyl acetate and phosphotungstic acid. Preparations were first incubated at pH 6.6, gamma/2 0.3 and 0 degrees C to develop fibrinogen aggregates at concentrations approaching equilibrium levels. The fraction of fibrinogen present as aggregates, designated the product mass fraction (Pmf), was determined by gel filtration on Bio-Gel A-1.5m. Electron micrographs of preparations with significant Pmf revealed partially closed quasi-globular structures of variable size. Fibrinogen aggregates could be dissociated by incubating preparations at 2 mg/ml, 37 degrees C for 18h, with a resulting Pmf less than 0.01. In these dissociated systems, monomers were observed with no apparent aggregate forms. At intervals during the approach to equilibrium, the size distributions of aggregates present were estimated by Bio-Gel A-15m filtration. A thermodynamic analysis incorporating these and previous data led to the following model specifications. Fibrinogen aggregates develop by monomer addition, followed by periodic reactions among the constituent members of an aggregate. These intra-aggregate reactions produce a non-linear structure, most likely begin after the association of four monomers, and on the average then follow the addition of every other monomer. In this manner, quasiglobular aggregates containing up to about 28 constituent monomers may develop as equilibrium is approached.