Synthesis and Characterization of Thrombin Conjugated γ‐Fe₂O₃ Magnetic Nanoparticles for Hemostasis

Abstract

Thrombin is the final protease produced in the clotting pathways. Thrombin has been used in the clinic more than six decades for topical hemostasis and wound management. In human plasma the half‐life of thrombin is shorter than 15 seconds due to close control by inhibitors. In order to stabilize thrombin, this enzyme was conjugated covalently and physically to γ‐Fe2O3 magnetic nanoparticles. The physical conjugation was accomplished through adsorption of thrombin to BSA coating on the nanoparticles. The coagulant activity of the covalently bound thrombin was significantly lower than that of the physically adsorbed thrombin. Leakage of the physically bound thrombin into PBS containing 4% HSA was negligible. The physical conjugation of thrombin onto the nanoparticles stabilized the thrombin against its major inhibitor antithrombin III and improved its storage stability. At optimal CaCl2 concentration, the clotting time by the bound thrombin is shorter than that of the free enzyme. This novel conjugated thrombin may be an efficient candidate for topical hemostasis and wound healing.

Novel magnetic fibrin hydrogel scaffolds containing thrombin and growth factors conjugated iron oxide nanoparticles for tissue engineering

“…9 Similarly, the leakage of both the covalently and the physically bound bFGF into PBS containing 4% HSA was not detected by the bFGF ELISA kit.…”

“…9,42 In the present study, these nanoparticles were used for the fabrication of the magnetic fibrin hydrogel scaffolds. The fibrin hydrogel scaffolds were prepared by mixing thrombinconjugated nanoparticles dispersed in an aqueous solution with a fibrinogen aqueous solution containing CaCl 2 , Factor XIII, aprotinin, growth factor-conjugated γ-Fe 2 O 3 nanoparticles, and the desired cells for cultivation.…”

“…9,42 Briefly, 5 mg of BSA or HSA were added to 4 mL of the γ-Fe 2 O 3 nanoparticles dispersed in water (2.5 mg/mL). The reaction mixture was subsequently shaken at pH 9.5 and 60°C for 18 hours, and then cooled gradually to room temperature.…”

“…7 In previous studies, we have demonstrated that appropriate conjugation of thrombin to iron oxide (γ-Fe 2 O 3 ) magnetic nanoparticles preserved the thrombin-clotting activity, stabilized the thrombin against its major inhibitor, antithrombin III, and improved its storage stability. 8,9 In the present study, these thrombin-conjugated γ-Fe 2 O 3 nanoparticles were used to fabricate novel magnetic fibrin hydrogel scaffolds.…”

Design of near-infrared fluorescent bioactive conjugated functional iron oxide nanoparticles for optical detection of colon cancer

“…25,31 Briefly, 20 mg of HSA and 2 mg of CANIR dissolved in 2 mL of PBS were added to 10 mL of fluorescent NIR IO nanoparticles dispersed in PBS (4 mg/mL, pH 7.4). The shaken mixture was gradually brought to 65°C, and after 16 hours at 65°C was cooled to room temperature.…”