Highly Sensitive Fluorescence-Linked Immunosorbent Assay for the Determination of Human IgG in Serum Using Quantum Dot Nanobeads and Magnetic Fe₃O₄ Nanospheres

Abstract

The aim of this study is to establish a new method with high sensitivity, accuracy, and stability for the determination of human IgG and then expand it to analyze severe acute respiratory syndrome corona virus 2 (SARS-CoV-2)-specific IgM and IgG, which is of great significance for the screening and diagnosis of COVID-19. In this study, the magnetic Fe₃O₄ nanospheres coupled with mouse antihuman IgG (Ab1_IgG) were used as an immune capture probe (Fe₃O₄@Ab1_IgG) to capture and separate the target, and rabbit antihuman IgG (Ab2_IgG) coupled with highly luminescent quantum dot nanobeads (QBs) as a fluorescence detection probe (QBs@Ab2_IgG) was used to realize high sensitivity detection. After the formation of a sandwich immunocomplex, the fluorescence intensity of the precipitate after magnetic separation was measured at the excitation wavelength of 370 nm. Under optimal conditions, a wide linear range varying from 0.005 to 40 ng·mL^-1 was obtained for the detection of human IgG with a lower limit of detection at 4 pg·mL^-1 (S/N = 3). The recoveries of intra- and interassays were 90.0-101.9 and 96.0-106.6%, respectively, and the relative standard deviations were 6.3-10.2 and 2.6-10.5%, respectively. Furthermore, the proposed method was successfully demonstrated to detect human IgG in serum samples, and the detection results were not statistically different (<i>P</i> > 0.05) from commercial enzyme-linked immunosorbent assay kits. This method is sensitive, fast, and accurate, which could be expanded to detect the specific IgM and IgG antibodies against SARS-CoV-2.