Ligand density characterization of peptide-modified biomaterials

被引:32
作者
Barber, TA
Harbers, GM
Park, S
Gilbert, M
Healy, KE
机构
[1] Univ Calif Berkeley, Dept Mat Sci & Engn, Berkeley, CA 94720 USA
[2] Northwestern Univ, Dept Biomed Engn, Robert R McCormick Sch Engn & Appl Sci, Evanston, IL 60208 USA
[3] Univ Calif Berkeley, Dept Bioengn, Berkeley, CA 94720 USA
关键词
peptide; surface analysis; surface modification; IPN; silane;
D O I
10.1016/j.biomaterials.2005.04.043
中图分类号
R318 [生物医学工程];
学科分类号
0831 ;
摘要
A simple fluorescence based characterization method was developed to assess ligand density on peptide-modified biomaterials. The method exploits the exquisite sensitivity of proteolysis for the purpose of liberating a fluorescently labeled probe fragment from an immobilized peptide. The released fragment can then be detected in solution using high-throughput fluorometry. In silico screening tools identified the enzyme chymotrypsin as a promising candidate for releasing a detectable probe fragment from the fluorescently labeled peptide, Ac-CGGNGEPRGDTYRAYK(FITC)GG-NH2. After chymotrypsin digestion of the peptide in solution was first characterized using mass spectrometry and HPLC, a basic enzyme mediated release protocol was developed and implemented to generate peptide-binding isotherms on various peptide-modified biomaterials. The new method is sensitive, has good signal-to-noise ratio (S/N), and is easily standardized. Furthermore, the technique can be applied independent of material chemistry and geometry, making it a suitable alternative to radiolabeling for a wide range of biomaterial applications. (c) 2005 Elsevier Ltd. All rights reserved.
引用
收藏
页码:6897 / 6905
页数:9
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