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

In certain conditions, dye-conjugated icosahedral virus shells exhibit suppression of concentration quenching. The recently observed radiation brightening at high fluorophore densities has been attributed to coherent emission, i.e., to a cooperative process occurring within a subset of the virussupported fluorophores. Until now, the distribution of fluorophores among potential conjugation sites and the nature of the active subset remained unknown. With the help of mass spectrometry and molecular dynamics simulations, we found which conjugation sites in the brome mosaic virus capsid are accessible to fluorophores. Reactive external surface lysines but also those at the lumenal interface where the coat protein N-termini are located showed virtually unrestricted access to dyes. The third type of labeled lysines was situated at the intercapsomeric interfaces. Through limited proteolysis of flexible N-termini, it was determined that dyes bound to them are unlikely to be involved in the radiation brightening effect. At the same time, specific labeling of genetically inserted cysteines on the exterior capsid surface alone did not lead to radiation brightening. The results suggest that lysines situated within the more rigid structural part of the coat protein provide the chemical environments conducive to radiation brightening, and we discuss some of the characteristics of these environments.

Radiation brightening was recently observed in a multifluorophoreconjugated brome mosaic virus (BMV) particle at room temperature under pulsed excitation. On the basis of its nonlinear dependence on the number of chromophores, the origins of the phenomenon were attributed to a collective relaxation. However, the mechanism remains unknown. We present ultrafast transient absorption and fluorescence spectroscopic studies which shed new light on the collective nature of the relaxation dynamics in such radiation-brightened, multifluorophore particles. Our findings indicate that the emission dynamics is consistent with a superradiance mechanism. The ratio between the rates of competing radiative and nonradiative relaxation pathways depends on the number of chromophores per virus. The findings suggest that small icosahedral virus shells provide a unique biological scaffold for developing nonclassical, deep subwavelength light sources and may open new avenues for the development of photonic probes for medical imaging applications.

Radiation brightening was recently observed in a multi-fluorophore-conjugated brome mosaic virus (BMV) particle, at room temperature under pulsed excitation. Based on its nonlinear dependence on the number of fluorophores, the origins of the phenomenon were attributed to a collective relaxation. However, the mechanism remains unknown.We present ultrafast transient absorption and fluorescence spectroscopic studies which shed new light on the collective nature of the relaxation dynamics in such radiationbrightened, multi-fluorophore particles. Our findings indicate that the emission dynamics is consistent with a superradiance mechanism. The ratio between the rates of competing radiative and non-radiative relaxation pathways depends on the number of fluorophores per virus. We also discuss the evidence of coherent oscillations in the transient absorption trace from multi-fluorophore conjugated which last for ∼ 100s of

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