Fluoroimmunoassay for tumor necrosis factor- alpha in human serum using Ru(bpy) sub(3)Cl sub(2)-doped fluorescent silica nanoparticles as labels

• Published in: Talanta (Oxford) Vol. 73; no. 2; pp. 366 - 371
• Main Authors: Hun, Xu, Zhang, Zhujun
• Format: Journal Article
• Language: English
• Published: 01.09.2007
• ISSN: 0039-9140
• DOI: 10.1016/j.talanta.2007.03.059

A novel fluoroimmunoassay (FIA) method was developed for the determination of tumor necrosis factor- alpha (TNF- alpha ) in this study. The proposed method has the advantage of showing the specificity of immunoassays and sensitivity of fluorescent nanoparticles label technology. With the well- established inverse microemulsion polymerisation process, the tris(2',2- bipyridyl)dichlororuthenium(II) hexahydrate (Rubpy)-doped fluorescent silica nanoparticles (RuDFSNs) were prepared. Then a RuDFSNs-labeled anti-TNF- alpha monoclonal antibody was prepared and used for FIA of TNF- alpha in human serum samples with a sandwich FIA by using the low fluorescent 96-well transparent microtiter plates. The assay response was linear from 1.0 to about 250.0 pg/mL with a detection limit of 0.1 pg/mL for TNF- alpha . The intra- and inter- assay precision are 4.9%, 4.4%, 4.6%; 6.1%, 5.9%, 5.3% for five parallel measurements of 2.0, 20.0, 200.0 pg/mL TNF- alpha respectively, and the recoveries are in the range of 96-104% for human serum sample measurements by standard-addition method. We also explored the application of fluorescence microscopy imaging in the study of the FIA for TNF- alpha with the fluorescent nanoparticle labels. The results demonstrate that the method offers potential advantages of sensitivity, simplicity and good reproducibility for the determination of TNF- alpha , and is applicable to the determination of TNF- alpha in serum samples and being capable of fluorescence microscopy imaging for the determination of TNF- alpha .