Decoration of Reduced Graphene Oxide Nanosheets with Aryldiazonium Salts and Gold Nanoparticles toward a Label-Free Amperometric Immunosensor for Detecting Cytokine Tumor Necrosis Factor-α in Live Cells.
In this study, a label-free electrochemical immunosensor was developed for detection of cytokine tumor necrosis factor-alpha (TNF-α). First, AuNPs loaded reduced graphene oxides nanocomposites (RGO-ph-AuNP) were prepared, and then, a mixed layer of 4-carbxyphenyl and 4-aminophenyl phosphorylcholine (PPC) was modified to the surface of AuNPs for the subsequent modification of anti-TNF-α capture antibody (Ab1) to form the capture surface (Au-RGO-ph-AuNP-ph-PPC(-ph-COOH)) for the analyte TNF-α with the antifouling property. For reporting the presence of analyte, the anti-TNF-α detection antibody (Ab2) was modified to the graphene oxides which have been modified with the 4-ferrocenylaniline through diazonium chemistry to form Ab2-GO-ph-Fc. Then, a sandwich assay was formed on gold surfaces for the quantitative detection of TNF-α based on the electrochemical signal of ferrocene. X-ray photoelectron spectra (XPS), transmission electron microscopy (TEM), Fourier transform infrared spectroscopy (FT-IR), UV-vis, and electrochemistry were used for characterization of the stepwise fabrications on the interface. The prepared electrochemical immunosensor was successfully used for the detection of TNF-α over the range of 0.1-150 pg mL-1. The lowest detection limit of this immunosensor is 0.1 pg mL-1TNF-α in 50 mM phosphate buffer at pH 7.0. The fabricated immunosensor provided high selectivity and stability and can be used to detect TNF-α secreted by live BV-2 cells with comparable accuracy to enzyme-linked immunosorbent assay (ELISA) but with lower limit of detection.
Key Laboratory of Pesticide and Chemical Biology of Ministry of Education, College of Chemistry, Central China Normal University , Wuhan, Hubei 430079, P. R. China.
Meng
M
Qi
Key Laboratory of Pesticide and Chemical Biology of Ministry of Education, College of Chemistry, Central China Normal University , Wuhan, Hubei 430079, P. R. China.
Yin
Y
Zhang
Key Laboratory of Pesticide and Chemical Biology of Ministry of Education, College of Chemistry, Central China Normal University , Wuhan, Hubei 430079, P. R. China.
Chaomin
C
Cao
Key Laboratory of Pesticide and Chemical Biology of Ministry of Education, College of Chemistry, Central China Normal University , Wuhan, Hubei 430079, P. R. China.
Mingxing
M
Zhang
Key Laboratory of Pesticide and Chemical Biology of Ministry of Education, College of Chemistry, Central China Normal University , Wuhan, Hubei 430079, P. R. China.
Shenghua
S
Liu
Key Laboratory of Pesticide and Chemical Biology of Ministry of Education, College of Chemistry, Central China Normal University , Wuhan, Hubei 430079, P. R. China.|ARC Centre of Excellence in Nanoscale Biophotonics (CNBP), Macquarie University , North Ryde, New South Wales 2109, Australia.
Guozhen
G
Liu
D000818:Animals; D015374:Biosensing Techniques; D002470:Cell Survival; D002478:Cells, Cultured; D003979:Diazonium Compounds / Q000737:chemistry*; D055664:Electrochemical Techniques; D006046:Gold / Q000737:chemistry; D006108:Graphite / Q000737:chemistry*; D007118:Immunoassay; D051379:Mice; D049329:Nanostructures / Q000737:chemistry*; D010084:Oxidation-Reduction; D010087:Oxides / Q000737:chemistry*; D012492:Salts / Q000737:chemistry; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis*
D003979:Diazonium Compounds; D010087:Oxides; D012492:Salts; D014409:Tumor Necrosis Factor-alpha; D006046:Gold; D006108:Graphite