Wu Yu-Wei, Chen Sung-Fang, Yang Charng-Bin, Tsai Yu-Hui
Graduate Institute of Pharmacy, Taipei Medical University, Taipei, Taiwan.
J Chromatogr B Analyt Technol Biomed Life Sci. 2008 Feb 15;863(1):187-91. doi: 10.1016/j.jchromb.2008.01.019. Epub 2008 Jan 18.
In this study, a protein purified by fluorescein isothiocyanate (FITC)-affinity chromatography from human plasma was identified as albumin by MALDI-TOF-MS. Albumin was found to conjugate with FITC-labeled molecules through a copper-dependent reaction. The formation of this complex was confirmed by methods including a newly developed "charcoal-based fluorescence assay" (CFA), gel-filtration, affinity chromatography, and ultrafiltration. The binding was identified as disulfide bridge formation. This is the first to demonstrate that copper induces a covalent binding of FITC-labeled molecules with albumin. In addition, the developed CFA method facilitates the screening of small fluorescent dyes binding to macromolecules.
