Monitoring Thrombin Generation and Screening Anticoagulants through Pulse Laser-Induced Fragmentation of Biofunctional Nanogold on Cellulose Membranes

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• 作者：Yu-Jia Li ; Wei-Jane Chiu ; Binesh Unnikrishnan ; Chih-Ching Huang
• 刊名：ACS Applied Materials & Interfaces
• 出版年：2014
• 出版时间：September 10, 2014
• 年：2014
• 卷：6
• 期：17
• 页码：15253-15261
• 全文大小：428K
• ISSN：1944-8252

文摘

Thrombin generation (TG) has an important part in the blood coagulation system, and monitoring TG is useful for diagnosing various health issues related to hypo-coagulability and hyper-coagulability. In this study, we constructed probes by using mixed cellulose ester membranes (MCEMs) modified with gold nanoparticles (Au NPs) for monitoring thrombin activity using laser desorption/ionization mass spectrometry (LDI鈥揗S). The LDI process produced Au cationic clusters ([Au_n]⁺; n = 1鈥?) that we detected through MS. When thrombin reacted with fibrinogen on the Au NPs鈥揗CEMs, insoluble fibrin was formed, hindering the formation of Au cationic clusters and, thereby, decreasing the intensity of their signals in the mass spectrum. Accordingly, we incorporated fibrinogen onto the Au NPs鈥揗CEMs to form Fib鈥揂u NPs鈥揗CEM probes to monitor TG with good selectivity (>1000-fold toward thrombin with respect to other proteins or enzymes) and sensitivity (limit of detection for thrombin of ca. 2.5 pM in human plasma samples). Our probe exhibited remarkable performance in monitoring the inhibition of thrombin activity by direct thrombin inhibitors. Analyses of real samples using our new membrane-based probe suggested that it will be highly useful in practical applications for the effective management of hemostatic complications.

Keywords:

thrombin generation; drugs screening; gold nanoparticles; cellulose membranes; laser desorption/ionization