title: Functional fusion proteins and prevention of electrode fouling for a sensitive electrochemical immunosensor.
abstract: A highly sensitive electrochemical immunosensor was developed by preventing electrode fouling and using a novel fusion protein of silica binding polypeptides (SBP)-protein G (ProG) created by recombinant DNA technology as a functional crosslinker for rapid and self-oriented immobilization of antibodies onto silica nanoparticles (SiNPs). Antibody immobilization onto the SiNPs by the SBP-ProG could rapidly be achieved without any chemical treatment. The immunosensor was fabricated through bonding of a partially gold-deposited cyclic olefin copolymer (COC) (top substrate) and gold patterned interdigitated array COC electrode (bottom substrate). To prevent electrode fouling, human immunoglobulin G (hIgG) was immobilized onto the ceiling inside the microchannel, instead of the bottom electrode. Alkaline phosphatase (AP)-labeled anti-hIgG was allowed to immunoreact with hIgG on the ceiling, followed by addition of an enzyme to generate an oxidative peak current. A three-fold increase in current was observed from the immunosensor without any electrode fouling compared with a control with the protein functionalized electrode. Also, the SiNPs facilely coated with AP-anti-hIgG via the SBP-ProG could increase the electrochemical signal up to 20% larger than that of the AP-anti-hIgG alone. Furthermore, this immunosensor was ultrasensitive with a detection limit of 0.68 pg/mL of a biomarker associated with prostate cancer.
authors.affiliation: Department of Biotechnology, CHA University, 335 Pangyo-ro, Bundang-gu, Seongnam, Gyeonggi-Do 13488, Republic of Korea.
authors.forename: A-Ram
authors.initials: AR
authors.lastname: Kim
authors.affiliation: Department of Chemistry, Chung-Ang University, 84 Heukseok-ro, Dongjak-gu, Seoul 06974, Republic of Korea.
authors.forename: Tae Jung
authors.initials: TJ
authors.lastname: Park
authors.affiliation: Department of New Biology, DGIST, 333, Techno Jungang-Daero, Daegu, 42988goo, Republic of Korea.
authors.forename: Minseok S
authors.initials: MS
authors.lastname: Kim
authors.affiliation: Korea Food Research Institute (KFRI), Bundang-gu, Seongnam, Gyeonggi-Do 463-836, Republic of Korea.
authors.forename: In-Ho
authors.initials: IH
authors.lastname: Kim
authors.affiliation: Predictive Model Research Center, Korea Institute of Toxicology (KIT), Daejeon, Republic of Korea; Department of Human and Environmental Toxicology, University of Science and Technology, Daejeon, 34113, Republic of Korea.
authors.forename: Ki-Suk
authors.initials: KS
authors.lastname: Kim
authors.affiliation: Department of Biotechnology, CHA University, 335 Pangyo-ro, Bundang-gu, Seongnam, Gyeonggi-Do 13488, Republic of Korea.
authors.forename: Kwang Hoe
authors.initials: KH
authors.lastname: Chung
authors.affiliation: Department of Biotechnology, CHA University, 335 Pangyo-ro, Bundang-gu, Seongnam, Gyeonggi-Do 13488, Republic of Korea. Electronic address: shko7@cha.ac.kr.
authors.forename: Sungho
authors.initials: S
authors.lastname: Ko
mesh_terms: D014408:Biomarkers, Tumor / Q000378:metabolism; D015374:Biosensing Techniques; D055664:Electrochemical Techniques; D004566:Electrodes; D006046:Gold; D006801:Humans; D007118:Immunoassay; D008297:Male; D053758:Nanoparticles; D017430:Prostate-Specific Antigen / Q000097:blood; D011471:Prostatic Neoplasms / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry*
chemical_list: D014408:Biomarkers, Tumor; D011993:Recombinant Fusion Proteins; D006046:Gold; D017430:Prostate-Specific Antigen
keywords: Cyclic olefin copolymer; Electrochemical immunosensor; Electrode fouling; Protein G; Silica binding polypeptide; Silica nanoparticle