Determination of Human IgG by Solid Substrate‐Room Temperature Phosphorescence Immunoassay

Abstract

Silicon dioxide nanoparticle containing fluorescein isothiocyanate (FITC‐SiO₂) with the particle size of 20 nm was synthesized by solgel method. It can emit strong and stable solid substrate‐room temperature phosphorescence (SS‐RTP) signal on the substrate of polyamide membrane. It was found in the research that the specific reaction between goat‐antihuman IgG antibody labeled with FITC‐SiO₂ and human IgG (immunoglobulin) can be carried on the polyamide membrane quantitatively, the antigen‐antibody conjugate retained good phosphorescence properties, and the signal of phosphorescence was enhanced after the immunoassay reaction. Based on the facts above, a new method of solid substrate‐room temperature phosphorescence immunoassay (SS‐RTP‐IA) to determine human IgG was established. The linear range of this method is 0.080–20.0 pg human IgG spot⁻¹ (sample volume 0.40 µl spot⁻¹, and the corresponding concentration range of human IgG is 0.20–50.00 ng ml⁻¹). The regression equation of the working curve is ΔIp = 19.97 + 15.92 m _IgG (pg spot⁻¹), n = 6, correlation coefficient is 0.9997. The detection limit of this method calculated by 3 Sb/k is 0.015 pg spot^–1. Compared with the immunoassay using FITC to label goat‐antihuman IgG, the sensitivity was obviously improved. For the sample containing 0.20 (ng ml⁻¹) and 50.0 (ng ml^–1) human IgG, with a sample volume of 0.40 µl, the relative standard deviation (RSD) is 3.4% and 4.1% (n = 11), respectively. This indicated very good repeatability.

Keywords:

• Human IgG
• Luminescent nanoparticle as labeling reagent
• Solid substrate‐room temperature phosphorescence immunoassay
• Fluorescein isothiocyanate

Acknowledgments

This work was supported by the Chinese National Natural Science Foundation (Grant No. 20175011) and Fu–jian Province Natural Science Foundation (Grant No. C0110028).