Ultrabright Terbium Nanoparticles for FRET Biosensing and in Situ Imaging of Epidermal Growth Factor Receptors

• Published in: Chemistry : a European journal Vol. 26; no. 64; pp. 14602 - 14611
• Main Authors: Charpentier, Cyrille, Cifliku, Vjona, Goetz, Joan, Nonat, Aline, Cheignon, Clémence, Cardoso Dos Santos, Marcelina, Francés‐Soriano, Laura, Wong, Ka‐Leung, Charbonnière, Loïc J., Hildebrandt, Niko
• Format: Journal Article
• Language: English
• Published: Germany Wiley Subscription Services, Inc 17-11-2020; Wiley-VCH Verlag
• Subjects: Animals; Antennas; Antibodies; Binding; bio-labeling; Biomolecules; Biosensing Techniques; Biosensors; Biotechnology; Cell lines; Chemical compounds; Chemical Sciences; Chemistry; Conjugates; EGF Family of Proteins; Energy transfer; Epidermal growth factor; Epidermal growth factor receptors; ErbB Receptors - metabolism; Fluorescence; Fluorescence Resonance Energy Transfer; Fluorophores; FRET; Growth factor receptors; Growth factors; Intracellular signalling; lanthanide nanoparticles; Life Sciences; Ligands; luminescence microscopy; Medical imaging; Mice; Nanoparticles; NIH 3T3 Cells; Physics; Receptors; Streptavidin; Terbium; time-resolved detection; time-resolved detection; lanthanide nanoparticles; luminescence microscopy; FRET; bio-labeling
• ISSN: 0947-6539; 1521-3765
• DOI: 10.1002/chem.202002007

Lanthanide‐doped nanoparticles (LnNPs) have become an important class of fluorophores for advanced biosensing and bioimaging. LnNPs that are photosensitized by surface‐attached antenna ligands can possess exceptional brightness. However, their functional bioconjugation remains an important challenge for their translation into bioanalytical applications. To solve this problem, we designed a ligand that can be simultaneously applied as efficient light harvesting antenna for Tb surface ions and strong linker of biomolecules to the LnNPs surfaces. To demonstrate generic applicability of the photosensitized TbNP‐bioconjugates, we applied them in two prototypical applications for biosensing and bioimaging. First, in‐solution biorecognition was shown by time‐resolved Förster resonance energy transfer (FRET) between streptavidin‐functionalized TbNPs to biotinylated dyes (ATTO 610). Second, in situ detection of ligand–receptor binding on cells was accomplished with TbNP‐antibody (Matuzumab) conjugates that could specifically bind to transmembrane epidermal growth factor receptors (EGFR). High specificity and sensitivity were demonstrated by time‐gated imaging of EGFR on both strongly (A431) and weakly (HeLa and Cos7) EGFR‐expressing cell lines, whereas non‐expressing cell lines (NIH3T3) and EGFR‐passivated A431 cells did not show any signals. Despite the relatively large size of TbNP‐antibody conjugates, they could be internalized by A431 cells upon binding to extracellular EGFR, which showed their potential as bright and stable luminescence markers for intracellular signaling.
Trifunctional ligands were engineered for surface anchoring on lanthanide based nanoparticles, photosensitization of the lanthanides, and biological functionalization with streptavidin or antibodies. The efficiency of these ultrabright lanthanide nanobioconjugates was demonstrated by Förster resonance energy transfer (FRET) biosensing and selective imaging of epidermal growth factor receptors at the surface of cancer cells.