Screening, purification, and identification of a copper-dependent FITC-binding protein in human plasma: Albumin

Yu Wei Wu, Sung-Fang Chen, Charng Bin Yang, Yu Hui Tsai

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

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.

Original languageEnglish
Pages (from-to)187-191
Number of pages5
JournalJournal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
Volume863
Issue number1
DOIs
Publication statusPublished - 2008 Feb 15

Fingerprint

Fluorescein
Serum Albumin
Purification
Affinity chromatography
Copper
Albumins
Carrier Proteins
Screening
Charcoal
Affinity Chromatography
Assays
Fluorescence
Plasma (human)
Molecules
Matrix-Assisted Laser Desorption-Ionization Mass Spectrometry
Ultrafiltration
Macromolecules
Fluorescent Dyes
Disulfides
Gel Chromatography

Keywords

  • Affinity
  • Albumin
  • Copper
  • FITC
  • Purification

ASJC Scopus subject areas

  • Biochemistry

Cite this

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abstract = "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.",
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AU - Chen, Sung-Fang

AU - Yang, Charng Bin

AU - Tsai, Yu Hui

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AB - 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.

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