The rapid diagnostic kit such as a pregnancy test is widely used. Samples of urine, serum and plasma are usually diluted with diluent buffer and separated by an immunochromatographic method. The analytes in the test can be immobilized to the nitrocellulose membrane which was coated with specific antibody or antigen and visualized by specific Ag or Ab conjugated with gold nanoparticles. However, their application is limited to the quantitative assay due to the Hook effect, which shows the false negative result in case of high concentrations of analyte in the sample.
Methods
To overcome the Hook effect, we devised an innovative rapid kit which has a unique competition method as well as a classical sandwich method in a single kit. In this study, test line 1 was prepared with mouse monoclonal anti-human IgG or anti-human IgE like classical rapid kits. Test line 2, a competitive line, which was coated with human IgG or human IgE, was designed to improve the false negative result due to the Hook effect. The control line was prepared with goat anti-mouse IgG. Goat anti-human IgG antibody and mouse IgG conjugated with gold nanoparticles were prepared in our laboratory and placed in the conjugate pad for further test.
Results
Immunecheck human IgG (Proteometech Inc, Seoul, Korea) can detect IgG range from 1.2 to 40,000 ug/ml. To know the degree of agreement against other quantitative assay method, we compared our assay results of 30 human sera with an immunoturbidimetric assay using Cobas Integra 800 (Roche, Indianapolis, IN, USA). This comparison showed that Immunecheck human IgG had 74.2 – 132.1% of degree of agreement and 0.9243 of coefficient of determination (R2) against an immunoturbidimetric method. The detection limit of Immunecheck human IgE was 80 IU/ml of total IgE and their signals were increased depending on the concentrations of IgE.
Conclusions
We developed a novel and innovative rapid immunoassay system which can overcome the hurdle of hook effect and detect the wide range of target molecule. This system will be beneficial to reduce the misleading of test results which cannot be possible with a current conventional immunoassay system.