A Peptide Microarray-Based Fluorescent and Resonance Light Scattering Assay for Screening Thrombin Inhibitor
10.11895/j.issn.0253-3820.140532
- VernacularTitle:基于多肽微阵列芯片的荧光和共振光散射法筛选凝血酶抑制剂
- Author:
Min SU
;
Tao LI
;
Dianjun LIU
;
Zhenxin WANG
- Publication Type:Journal Article
- Keywords:
Peptide microarray;
Gold nanoparticle probes;
Resonance light scattering;
Half maximal inhibitory concentration;
Thrombin inhibitor
- From:
Chinese Journal of Analytical Chemistry
2015;(2):199-206
- CountryChina
- Language:Chinese
-
Abstract:
A peptide microarray-based fluorescence and resonance light scattering ( RLS ) two readout assay was developed for screening thrombin inhibitors in blood samples. In this assay, the biotinylated peptide microarray was used as the platform. The peptide C-terminal fragments carried biotin sites departed from the slide when the biotinylated peptides were digested by thrombin hydrolysis reaction. The hydrolysis progress was labeled by fluorescence and 30 nm peptide-stabilized gold nanoparticles through the biotin-avidin reaction. In the presence of thrombin inhibitors, the hydrolysis reactions were blocked, and the inhibition capability of inhibitors could be detected by the fluorescent and RLS signal changes. The order of the half maximal inhibitory concentration ( IC50 ) of thrombin inhibitors in pure thrombin solution and spiked human serum were argatrobanHAT-Ⅲ>trypsin inhibitor>E-64>AEBSF. The reversible or irreversible characters of argatroban and HAT-Ⅲ had been estimated in human plasma. Compared with the experimental data of fluorescent and RLS assay in blood sample, the RLS assay labeled by 30 nm gold nanoparticles are more suitable for the inhibitor detection in complicated blood sample.
