RNA Isolation from Fibrin and Plasma

A mirVana RNA Isolation Kit (Thermo Fisher Scientific, Waltham, MA, USA) was used to isolate total RNA from fibrin. A mirVana PARIS Kit (Thermo Fisher Scientific, Waltham, MA, USA) was used to isolate RNA from 200 μL of plasma according to the manufacturer’s instructions. Briefly, recommended volume of miRNA homogenate additive (mirVana,) or 2X Denaturing Solution (PARIS) was added to each sample and mixed by vortexing. After adding 2X Denaturing Solution, plasma samples were spiked with synthetic cel-39 and incubated on ice. An equal volume of acid-phenol: and chloroform (Ambion) was added to each aliquot. The upper aqueous phase solutions were separated and mixed with 1.25 volumes of 100% molecular-grade ethanol. The solution was passed through a mirVana or PARIS column according to the manufacturer’s instructions. Finally, 100 µL of preheated 95 °C elution solution was used to elute total RNA. The concentration of total RNA isolated from fibrin was quantified using a NanoDrop 2000c spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA) [32 (link),33 (link),34 (link)].