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

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

Methods Appl Fluoresc

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.

Published: January 2021

AI Article Synopsis

Article Abstract

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.

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Source
http://dx.doi.org/10.1088/2050-6120/abd8e4DOI Listing

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