Determination of human IgG by solid-substrate room-temperature phosphorescence immunoassay based on an antibody labeled with nanoparticles containing rhodamine 6G luminescent molecules

• Published in: Analytical and bioanalytical chemistry 2004-10, Vol.380 (4), p.632-636
• Main Authors: Liu, Jia-Ming, Yang, Tian-Long, Liang, Xue-Shui, Wu, Ai-Hong, Li, Long-Di, Lin, Shao-Qin
• Format: Article
• Language: English
• Subjects: Animals; Antibodies - analysis; Antibodies - immunology; Antigen-Antibody Complex - analysis; Antigen-Antibody Complex - immunology; Humans; Immunoassay - instrumentation; Immunoassay - methods; Immunoglobulin G - analysis; Immunoglobulin G - blood; Immunoglobulin G - immunology; Indicator Dilution Techniques; Luminescent Measurements; Microscopy, Electron, Transmission; Nanotechnology - methods; Rhodamines - analysis; Rhodamines - chemistry; Sensitivity and Specificity; Silicon Dioxide - chemistry; Spectrum Analysis; Temperature; Time Factors
• ISSN: 1618-2642; 1618-2650
• DOI: 10.1007/s00216-004-2767-y

Luminescent 50-nm silicon dioxide nanoparticles containing both types of rhodamine 6G (R; particles denoted R-SiO2) were synthesized by the sol-gel method. In the presence of Pb(Ac)2 as a heavy atom perturber the particle can emit the intense and stable room-temperature phosphorescence (RTP) signal of R on a polyamide membrane, with lambda(ex)max/ lambda(em)max=470/635 nm for R. Our research indicates that the specific immune reaction between goat-anti-human IgG antibody labeled with R-SiO2 and human IgG can be carried out quantitatively on a polyamide membrane, and the phosphorescence intensity was enhanced after the immunoreaction. Thus a new method for solid-substrate room-temperature phosphorescence immunoassay (SS-RTP-IA) for determination of human IgG was established on the basis of antibody labeled with the nanoparticles containing binary luminescent molecules. The linear range of this method is 0.0624-20.0 pg spot(-1) of human IgG (corresponding to a concentration range of 0.156-50.0 ng mL(-1), sample volume 0.40 microL spot(-1)). The regression equations of the working curves are DeltaIp = 71.27+7.208 m(IgG) (pg spot(-1)) (r = 0.9996). Detection limits calculated as 3 Sb/k are 0.022 pg spot(-1). Compared with the same IA using fluorescein isothiocyanate (FITC) as the marker the new method was more sensitive and had a wider linear range. After elevenfold replicate measurement RSD are 4.5 and 3.6% for samples containing 0.156 and 50.0 ng mL(-1) IgG, respectively. This method is sensitive, accurate, and of high precision.