Optimizing fluorophore density for single virus counting: a photophysical approach.

Autor:	Chatterjee S; Nanobiophysics (NBP), MESA + Institute for Nanotechnology and Technical Medical Centre, Faculty of Science and Technology, University of Twente, PO Box 217, 7500 AE Enschede, The Netherlands.; Wetsus, European Centre of Excellence for Sustainable Water Technology, Oostergoweg 9, 8911MA, Leeuwarden, The Netherlands., Molenaar R; Nanobiophysics (NBP), MESA + Institute for Nanotechnology and Technical Medical Centre, Faculty of Science and Technology, University of Twente, PO Box 217, 7500 AE Enschede, The Netherlands., Tromp L; Nanobiophysics (NBP), MESA + Institute for Nanotechnology and Technical Medical Centre, Faculty of Science and Technology, University of Twente, PO Box 217, 7500 AE Enschede, The Netherlands., Wagterveld RM; Wetsus, European Centre of Excellence for Sustainable Water Technology, Oostergoweg 9, 8911MA, Leeuwarden, The Netherlands., Roesink HDW; Membrane Science & Technology (MST), MESA + Institute for Nanotechnology, Faculty of Science and Technology, University of Twente, PO Box 217, 7500 AE Enschede, The Netherlands., Cornelissen JJLM; Biomolecular Nanotechnology (BNT), MESA + Institute for Nanotechnology, Faculty of Science and Technology, University of Twente, PO Box 217, 7500 AE Enschede, The Netherlands., Claessens MMAE; Nanobiophysics (NBP), MESA + Institute for Nanotechnology and Technical Medical Centre, Faculty of Science and Technology, University of Twente, PO Box 217, 7500 AE Enschede, The Netherlands., Blum C; Nanobiophysics (NBP), MESA + Institute for Nanotechnology and Technical Medical Centre, Faculty of Science and Technology, University of Twente, PO Box 217, 7500 AE Enschede, The Netherlands.
Jazyk:	angličtina
Zdroj:	Methods and applications in fluorescence [Methods Appl Fluoresc] 2021 Jan 22; Vol. 9 (2), pp. 025001. Date of Electronic Publication: 2021 Jan 22.
DOI:	10.1088/2050-6120/abd8e4
Abstrakt:	In health and environmental research, it is often necessary to quantify the concentrations of single (bio) nanoparticles present at very low concentrations. Suitable quantification approaches that rely on counting and tracking of single fluorescently labelled (bio) nanoparticles are often challenging since fluorophore self-quenching limits the maximum particle brightness. Here we study how the number of labels per nanoparticle influences the total brightness of fluorescently labelled cowpea chlorotic mottle virus (CCMV). We analyze in detail the photophysical interplay between the fluorophores on the virus particles. We deduce that the formation of dark aggregates and energy transfer towards these aggregates limits the total particle brightness that can be achieved. We show that by carefully selecting the number of fluorescent labels per CCMV, and thus minimizing the negative effects on particle brightness, it is possible to quantify fluorescently labelled CCMV concentrations down to fM concentrations in single particle counting experiments.
Databáze:	MEDLINE
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