<p>(A) Effects of longistatin on the formation of fibrin clot. Fibrinogen (7.5 mM) was pre-incubated in a buffer in the absence or presence of longistatin (0.1, 0.2, 0.4, 0.8 and 1.6 µM) or plasmin (1.6 µM) and then thrombin was added (0.10 NIH unit/µl) as described in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1001312#s4" target="_blank">Materials and Methods</a>. Clot formation was detected visually and also by determining changes in turbidity at OD<sub>450</sub> using a spectrophotometer after 15 min. (B) Longistatin (1.6 µM) delayed fibrin clot formation up to 90 min. Fibrinogen was incubated in the absence or presence of longistatin (1.6 µM) following the same procedures as mentioned in A and then thrombin was added. OD<sub>450</sub> was measured at 15 min intervals. (C) Fibrinogenolytic effect of longistatin. Fibrinogen (7.5 mM) was incubated in the absence or presence of longistatin (0.4, 0.8 and 1.6 µM) or plasmin (1.6 µM). Samples were collected at the indicated time period and were subjected to 12.5% SDS–PAGE analysis under reducing conditions. A gradual degradation of the α, β and γ chains of fibrinogen was detectable with the concomitant deposition of degraded products. Asterisks (<sup>*</sup>) indicate that the difference compared with the negative control group (buffer only) is significant as determined by Student's <i>t</i>-test with unequal variance (<sup>*</sup>p<0.05, <sup>**</sup>p<0.01).</p
