SDS-PAGE analysis of soluble Tokyo V fibrin. Tokyo V fibrinogen was incubated with thrombin (1 U/mL) and FXIII (1 U/mL) in the presence of 2 mM calcium (Ca2+) or 2 mM EDTA (EDTA) at 37°C for 1 hour. Formed clots were squeezed using bamboo sticks to separate soluble fibrin (sFbn) from fibrin clots. (A) Untreated fibrinogen, insoluble clots, and soluble fractions were analyzed by SDS-PAGE using 7.5% polyacrylamide gels under nonreducing and reducing conditions. (B) For analysis of cross-linked soluble Tokyo V fibrin, the supernatant of the patient fibrin preparation was separated on 3% polyacrylamide gels under nonreducing conditions. Fibrinogen (Fbg) was the control. The reduced supernatant sample was analyzed by Western blotting for identification of γ chain. γ-dimers (γ-γ) and the aberrant γ chain (aberrant γ) were indicated.
