Genetically functionalized ferritin nanoparticles with a high-affinity protein binder for immunoassay and imaging

다국어 초록 (Multilingual Abstract)

<P><B>Abstract</B></P> <P>Molecular detection of target molecules with high sensitivity and specificity is of great significance in bio and medical sciences. Here, we present genetically functionalized ferritin nanoparticles with a high-affinity protein binder, and their utility as a signal generator in a variety of immunoassays and imaging. As a high-affinity protein binder, human IgG-specific repebody, which is composed of LRR (Leucine-rich repeat) modules, was used. The repebody was genetically fused to the N-terminal heavy-chain ferritin, and the resulting subunits were self-assembled to the repebody-ferritin nanoparticles composed of 24 subunits. The repebody-ferritin nanoparticles were shown to have a three-order of magnitude higher binding affinity toward human IgG than free repebody mainly owing to a decreased dissociation rate constant. The repebody-ferritin nanoparticles were conjugated with fluorescent dyes, and the resulting nanoparticles were used for western blotting, cell imaging, and flow cytometric analysis. The dye-labeled repebody-ferritin nanoparticles were shown to generate about 3-fold stronger fluorescent signals in immunoassays than monovalent repebody. The repebody-functionalized ferritin nanoparticles can be effectively used for sensitive and specific immunoassays and imaging in many areas.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Human IgG-specific repebody was genetically fused to N-terminus of ferritin subunit. </LI> <LI> The RbF-NPs exhibited 1000-fold higher binding affinity for human IgG than free repebody due to multivalency. </LI> <LI> The Cy3-laleled RbF-NPs generated stronger fluorescent signals than Cy3-labeled free repebody in immunoassays and imaging. </LI> <LI> The RbF-NPs can be effectively used in various types of immunoassays and imaging. </LI> </UL> </P> <P><B>Graphical abstract</B></P> <P>[DISPLAY OMISSION]</P>