Radiation Brightening from Virus-like Particles

Abstract

Concentration quenching is a well-known challenge in many fluorescence imaging applications. Here we show that the optical emission from hundreds of chromophores confined onto the surface of a virus particle 28 nm diameter can be recovered under pulsed irradiation. We have found that, as one increases the number of chromophores tightly-bound to the virus surface, fluorescence quenching ensues at first, but when the number of chromophores per particle is nearing the maximum number of surface sites allowable, a sudden brightening of the emitted light and a shortening of the excited state lifetime are observed. This radiation brightening occurs only under short pulse excitation; steady-state excitation is characterized by conventional concentration quenching for any number of chromophores per particle. The observed suppression of fluorescence quenching is consistent with efficient, collective radiation at room temperature. Interestingly, radiation brightening disappears when the emitters spatial and/or dynamic heterogeneity is increased, suggesting that the template structural properties may play a role and opening a way towards novel, virus-enabled imaging vectors that have qualitatively different optical properties than state-of-the-art biophotonic agents. 1 arXiv:1907.00065v2 [physics.app-ph] 2 Jul 2019 Abbreviations VLP Keywords virus nanotechnology, directed assembly, biophotonics, nanolaser, sub-wavelength, superradiance, quantum coherence, fluorescence quenching Photoluminescence, particularly fluorescence, is used in a myriad of applications in which low-background, high spatial resolution, and rapid response are required for non-intrusive imaging, remote sensing, and control of transient chemical states. Thus, synthetic fluorophores are incorporated in sensors and detectors, e.g. for early warning of bio-aerosolthreats, used in operation rooms for intraoperative guidance in brain and prostate cancer surgery, 1 and in anti-counterfeiting materials. 2 Over the years spectacular improvements in chromophore photostability, wavelength range, biological integration, detectors and detection techniques, have pushed the limits of fluorescence imaging to realms never thought possible before. 3,4 Despite these improvements, and somewhat surprisingly in the context of ever more demanding cutting edge applications, fluorescence emission is still overwhelmingly by way of uncorrelated, random emission from multiple chromophores. Associated with this regime are undesirable characteristics such as self-quenching, when emitters are too close, and exponential decay that is relatively slow at molecular scale (typically, 1-5 ns). Extended excited state lifetimes limit emission brightness, increasing the likelihood of photobleaching, and making the quantum yield more prone to change in response to environmental fluctuations. 5 At the same time, a system's collective behavior can be much more than the sum of its

Emergence by Design in Self-Assembling Protein Shells

“…20 Densely arranged fluorophores, when geometrically organised by site-specific conjugation to Brome mosaic virus capsids, can be coaxed into quantum coherence manifest as cooperative radiative decay. 21 However, and notwithstanding examples of polymorphism and tolerance to initial defects in natural protein shells, 13,22 a considerable drawback of adapting existing structures for new functions is the susceptibility of the system to defects in single components.…”

Ultrafast Collective Excited State Dynamics of a Virus-supported Fluorophore Antenna

“…The color scale corresponds to the difference between the absorbance of the samples in excited and ground-state (units of m∆A) complex is strongly accelerated with respect to that of the free dye. 11 The brightening effect was found to be a nonlinear function of N , the average number of fluorophores per particle.…”

“…In this work, a fluorescein-derived dye, Oregon Green TM 488, was covalently bound to the BMV capsid via NHS ester labelling of exposed lysines. 11 Maximum labeling density was ∼ 300 dyes/virus 11 (see Figure SI1). Figure 1A shows a molecular model of BMV, with the dye-accessible external and internal lysines colored in red and green respectively.…”

“…Radiation brightening from dye-conjugated fluorescent virus-like particles (fVLPs) was first reported by Tsvetkova et al . 11 It was found that when the fluorophores are conjugated with reactive residues of the brome mosaic virus (BMV) capsid interfaces, emission by the Figure 1: A) Schematic of accessible labelling sites on BMV capsid: red -external lysines K64,K105, K165 and green -internal lysines K41, K44 and K86. B) and C) transient absorption surface plots of free OG and BMV-OG278 following an initial excitation at 7.1 mJ/cm 2 .…”

“…The minimum nearest neighbour distance between lysines was estimated to be 1.9 nm. 11 To follow the excited state dynamics of coupled fluorophores in fVLPs, we performed pump-probe femtosecond, transient absorption (TA) spectroscopy. Differential absorption spectra of isolated fluorophores and fVLP samples with an average number of 278 dyes per particle (hereby called BMV-OG278) are presented as a function of the wavelength and timedelay in Figure 1B and Figure 1C, respectively.…”