Kim, Seong-Eun Ahn, Keum-Young Park, Jin-Seung Kim, Kyung Rim Lee, Kyung Eun Han, Sung-Sik Lee, Jeewon 2024-01-20T16:32:33Z 2024-01-20T16:32:33Z 2021-09-04 2011-08-01 0003-2700 https://pubs.kist.re.kr/handle/201004/130093 We synthesized fluorescent ferritin nanoparticles (FFNPs) through bacterial expression of the hybrid gene consisting of human ferritin heavy chain (hFTN-H), spacer (glycine-rich peptide), and enhanced green (or red) fluorescent protein [eGFP (or DsRed)] genes. The self-assembly activity of hFTN-H that leads to the formation of nanoparticles (12 nm in diameter), the conformational flexibility of the C-terminus of hFTN-H, and the glycine-rich spacer enabled eGFPs (or DsReds) to be well displayed on the surface of each ferritin nanoparticle, resulting in the construction of green (or red) FFNPs [gFFNPs (or rFFNPs)]. As compared to eGFP (or DsRed) alone, it is notable that the developed FFNPs showed significantly amplified fluorescence intensity and also enhanced stability. DNA aptamers were chemically conjugated to gFFNP via each eGFP's cysteine residue that was newly introduced through site-directed mutagenesis (Ser175Cys). The DNA-aptamer-conjugated gFFNPs were used as a fluorescent reporter probe in the aptamer-based "sandwich" assay of a cancer marker [i.e., platelet-derived growth factor B-chain homodimer (PDGF-BB)] in phosphate-buffered saline buffer or diluted human serum. This is a simple two-step assay without any additional steps for signal amplification, showing that compared to the same aptamer-based assays using eGFP alone or Cy3, the detection signals, affinity of the reporter probe to the cancer marker, and assay sensitivity were significantly enhanced; i.e., the limit of detection was lowered to the 100 fM level. Although the PDGF-BB assay is reported here as a proof-of-concept, the developed FFNPs can be applied in general to any aptamer-based sandwich assays. English AMER CHEMICAL SOC GROWTH-FACTOR QUANTUM DOTS PROTEIN NANOPARTICLES ESCHERICHIA-COLI ANTIBODY-BINDING GENE-EXPRESSION HIGH-AFFINITY H-CHAIN DNA MUTATIONS Fluorescent Ferritin Nanoparticles and Application to the Aptamer Sensor Article 10.1021/ac200657s 1 ANALYTICAL CHEMISTRY, v.83, no.15, pp.5834 - 5843 ANALYTICAL CHEMISTRY 83 15 5834 5843 scie scopus 000293252500007 2-s2.0-79961000174 Chemistry, Analytical Chemistry Article GROWTH-FACTOR QUANTUM DOTS PROTEIN NANOPARTICLES ESCHERICHIA-COLI ANTIBODY-BINDING GENE-EXPRESSION HIGH-AFFINITY H-CHAIN DNA MUTATIONS