Single-Protein Molecular Interactions on Polymer-Modified Glass Substrates for Nanoarray Chip Application Using Dual-Color TIRFM

• Published in: Bulletin of the Korean Chemical Society Vol. 28; no. 5; pp. 783 - 790
• Main Authors: Daekwang Kim, 이희구, 정형일, 강성호
• Format: Journal Article
• Language: English
• Published: 대한화학회 20.05.2007
• Subjects: 화학
• ISSN: 0253-2964; 1229-5949
• DOI: 10.5012/bkcs.2007.28.5.783

The immobilization of proteins and their molecular interactions on various polymer-modified glass substrates [i.e. 3-aminopropyltriethoxysilane (APTS), 3-glycidoxypropyltrimethoxysilane (GPTS), poly (ethylene glycol) diacrylate (PEG-DA), chitosan (CHI), glutaraldehyde (GA), 3-(trichlorosilyl)propyl methacrylate (TPM), 3'-mercaptopropyltrimethoxysilane (MPTMS), glycidyl methacrylate (GMA) and poly-l-lysine (PL).] for potential applications in a nanoarray protein chip at the single-molecule level was evaluated using prism-type dual-color total internal reflection fluorescence microscopy (dual-color TIRFM). A dual-color TIRF microscope, which contained two individual laser beams and a single high-sensitivity camera, was used for the rapid and simultaneous dual-color detection of the interactions and colocalization of different proteins labeled with different fluorescent dyes such as Alexa Fluor 488, Qdot 525 and Alexa Fluor 633. Most of the polymer-modified glass substrates showed good stability and a relative high signal-to-noise (S/N) ratio over a 40-day period after making the substrates. The GPTS/CHI/GA-modified glass substrate showed a 13.5-56.3% higher relative S/N ratio than the other substrates. 1% Top-Block in 10 mM phosphate buffered saline (pH 7.4) showed a 99.2% increase in the blocking effect of non-specific adsorption. These results show that dual-color TIRFM is a powerful methodology for detecting proteins at the single-molecule level with potential applications in nanoarray chips or nano-biosensors. KCI Citation Count: 14