Label-Free and Sensitive Detection of Thrombomodulin, a Marker of Endothelial Cell Injury, Using Quartz Crystal Microbalance.
Thrombomodulin (TM), an integral glycoprotein on the surface of endothelial cells, can be released during endothelial cell injury and the levels of serum TM are regarded as an important parameter of activity in vasculitides in vivo. Quantitative detection of TM and investigation on the release of soluble thrombomodulin (sTM) by the injured HUVEC-C cells using quartz crystal microbalance (QCM) were achieved in this work. Anti-antibody (AAb) and bovine serum albumin (BSA) were bound on gold nanoparticles (GNPs) to construct BSA-GNPs-AAb nanocomposites and they were characterized by transmission electron microscope, UV-vis, and infrared spectrophotometry, respectively. The capture of the nanocomposites on the TM antibody modified electrode, which was tested by scanning electron microscope, could result in a great decrease of the resonant frequency (f0). This binding was effectively inhibited by the beforehand immobilized TM proteins on the electrode surface due to the strong steric hindrance effect. It led to the decrease of the frequency changing extent. The relative frequency-shift was found to be proportional to the logarithm of the TM concentration from 10 to 5000 ng mL(-1) with a detection limit of 2 ng mL(-1). By analyzing the growth medium used for cell incubation, the release of sTM by the injured HUVEC-C cells in the presence of H2O2 was confirmed. The sTM amount in the growth medium was increased with the enhancement of contact time of the cells with H2O2, proving that sTM may serve as a specific marker of endothelial cell injury.
Key Laboratory of Chemical Biology and Traditional Chinese Medicine Research (Ministry of Education of China), College of Chemistry and Chemical Engineering, Hunan Normal University , Changsha, 410081, People's Republic of China.
Yiqun
Y
Luo
Key Laboratory of Chemical Biology and Traditional Chinese Medicine Research (Ministry of Education of China), College of Chemistry and Chemical Engineering, Hunan Normal University , Changsha, 410081, People's Republic of China.
Tong
T
Liu
Key Laboratory of Chemical Biology and Traditional Chinese Medicine Research (Ministry of Education of China), College of Chemistry and Chemical Engineering, Hunan Normal University , Changsha, 410081, People's Republic of China.
Jiaming
J
Zhu
Key Laboratory of Chemical Biology and Traditional Chinese Medicine Research (Ministry of Education of China), College of Chemistry and Chemical Engineering, Hunan Normal University , Changsha, 410081, People's Republic of China.
Liyan
L
Kong
Key Laboratory of Chemical Biology and Traditional Chinese Medicine Research (Ministry of Education of China), College of Chemistry and Chemical Engineering, Hunan Normal University , Changsha, 410081, People's Republic of China.
Wen
W
Wang
Key Laboratory of Chemical Biology and Traditional Chinese Medicine Research (Ministry of Education of China), College of Chemistry and Chemical Engineering, Hunan Normal University , Changsha, 410081, People's Republic of China.
Liang
L
Tan
D015415:Biomarkers / Q000032:analysis; D002478:Cells, Cultured; D004566:Electrodes; D042783:Endothelial Cells / Q000378:metabolism* / Q000473:pathology*; D006801:Humans; D010316:Particle Size; D058525:Quartz Crystal Microbalance Techniques; D013499:Surface Properties; D018180:Thrombomodulin / Q000032:analysis*
D015415:Biomarkers; D018180:Thrombomodulin