Analytical Sciences

Carbon Nanofiber-based Luminol-biotin Probe for Sensitive Chemiluminescence Detection of Protein

Stefan BAJ,* Tomasz KRAWCZYK,* Natalia PRADEL,* Md. Golam AZAM,** Takayuki SHIBATA,** Shpend DRAGUSHA,** Krzysztof SKUTIL,* Miroslawa PAWLYTA,*** and Masaaki KAI**

*Department of Chemical Organic Technology and Petrochemistry, Faculty of Chemistry, Silesian University of Technology, ul. Krzywoustego 4, 44-100 Gliwice, Poland
**Faculty of Pharmaceutical Sciences, Graduate School of Biomedical Sciences, Nagasaki University, 1-14 Bunkyo-machi, Nagasaki 852-8521, Japan
***Institute of Engineering Materials and Biomaterials, Silesian University of Technology, ul. Konarskiego 18a, 44-100 Gliwice, Poland

A carbon nanofiber-based luminol-biotin probe was synthesized for the sensitive chemiluminescence (CL) detection of a target protein by grafting luminol and biotin onto an oxidized carbon nanofiber. This carbon nanofiber was prepared by chemical vapor-deposition with methane in the presence of the Ni–Cu–MgO catalyst, which was followed by oxidization with HNO₃–H₂SO₄ to produce a carboxyl group on the surface of the nanofiber. The material was grafted with luminol and biotin by means of a standard carbodiimide activation of COOH groups to produce corresponding amides. The substance was water-soluble and thus could be utilized as a sensitive CL probe for a protein assay. The probe showed highly specific affinity towards the biotin-labeled antibody via a streptavidin–biotin interaction. The detection limit for this model assay was approximately 0.2 pmol of the biotinized IgG spotted on a polyvinylidene fluoride (PVDF) membrane. Nonspecific binding to other proteins was not observed. Therefore, the synthesized carbon nanofiber-based CL probe may be useful for a sensitive and specific analysis of the target protein.

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