Over the past years we have been applying quantum optical techniques to various biological problems such as: The detection of biopathogens, e.g., anthrax [1] and the SARS-CoV-2 virus [2]. On the theoretical front, I will discuss the application of quantum mechanics to other biological problems, such as the possible connection between quantum coherence in brain microtubules [3] and superradiance [4,5].

[1] “FAST CARS: Engineering a laser spectroscopic technique for rapid identification of bacterial spores,” M. Scully, et al., PNAS (2002).
[2] “Laser technique for direct identification of a single virus I: FASTER CARS,” V. Deckert, et al., PNAS (2020).
[3] “Quantum Coherence in (Brain) Microtubules and Efficient Energy and Information Transport”, N. Mavromatos and d. Nanopoulos, J. Phys: Conf. Ser. 329, 012026 (2011).
[4] “On the Existence of Superradiant Excitonic States in Microtubules”, G. Celardo, et al., NJP (2019).
[5] “The Super of Superradiance”, M. Scully and A. Svidzinksy, Science (2009): A. Svidzinsky, et al., “Single photon superradiance and radiation trapping by atomic shells,” PRA (2016).

Human exhaled breath contains more than 1,000 volatile organic molecules (biomarkers), combinations of which reflect metabolic processes in the human body. Spectroscopic detection of these biomarkers may be the basis for early diagnosis of various medical conditions. Our detection method, dual-comb spectroscopy (DCS) through the ultra-wideband, 1–100 THz (3–300 µm), spectral range, which targets the strongest vibrational and rotational molecular absorption resonances, relies on optical rectification of few-optical-cycle laser pulses to create mid-IR/THz frequency combs that are subsequently detected by electro-optical sampling (EOS), where the electric field of mid-IR/THz transients induces a change of polarization state of the near-IR probe pulse in the EO crystal. With this technique, we simultaneously achieved superior spectral resolution (via resolving comb lines), real-time detection, broadband (near octave) instantaneous coverage, and massive parallelism of data acquisition. Theoretically, EOS method is sensitive enough to detect vacuum fluctuations, which opens up wide opportunities for the study of quantum effects in biology.

This talk reflects a collaboration between Ye/Nesbitt groups to use of broad-band, ultrastable infrared frequency comb light sources in the 3-5 and 5-10 m IR fingerprint region with high finesse resonant cavities to probe small molecule content in exhaled human breath. These methods offer as much as a 100-million-fold enhancement in sensitivity x spectral throughput over conventional approaches, which in combination with machine learning algorithms have permitted encouraging first successes in real time identification of COVID disease state.

Enzymes are the workhorses of the cell and catalyze their reactions with enormous rate enhancements. The role of H-tunneling has been identified in a wide range of such reactions and shown to be dependent on a directed interplay between the extended protein scaffold and site of reactivity.

Weak magnetic fields affect a multitude of biological processes including cell metabolism and are hypothesized to be a result of magnetic field-sensitive spin-selective radical-pair reactions. To provide much needed visualization of this process, we demonstrate the use of a custom-built multimodal nonlinear optical imaging system capable of measuring the redox state of cells through multi-photon-excited autofluorescence and autofluorescence lifetime of metabolic cofactors. We demonstrate a custom multi-axis Helmholtz coil system to apply time-varying magnetic fields across the sample during imaging. This imaging platform allows for characterization and optimization of the effects of magnetic fields on live cells and tissues.

Small migratory songbirds are extraordinary navigators: weighing less than 30 g, they fly thousands of kilometres between their breeding and wintering grounds, alone and at night. To do so they use the sun and the stars, olfaction and landmarks, but it is clear that they can also perceive the direction of the Earth’s magnetic field. Despite more than 50 years of research, the biophysical mechanism of this remarkable magnetic sense remains obscure. In this talk, I will discuss the proposal that the birds’ magnetic compass relies on a quantum mechanism in their eyes. Specifically, the unique properties of light-induced radical pairs in cryptochrome proteins in photoreceptor cells could allow chemical sensing of magnetic interactions orders of magnitude weaker than might otherwise be thought possible.

Melanopsin, a tri-stable photopigment found in intrinsically-photosensitive retinal ganglion cells (ipRGCs), drives circadian rhythms and other non-image forming functions in the nervous system. Despite increased understanding of the biomolecular and spectroscopic properties of melanopsin, its multiphoton and ultrafast optical absorption properties remain underexplored. We demonstrate the effects of two-photon absorption of melanopsin using 900-1160 nm optical stimulation. Excitation in this bandwidth causes consistent increases in calcium levels in transfected HEK293T cells. Our results demonstrate the first reported nonlinear optical properties and corresponding functional responses of two-photon excitation of melanopsin in vitro, along with the effects of spectral-phase modulation on activation.

Previously we have found that the two-photon absorption (2PA) spectra of red fluorescent proteins have enhanced vibronic transitions and are about twofold broader, compared to the corresponding one-photon absorption (1PA) spectra. We explained these observations by the dependence of the permanent dipole moment change (Deltamu) on a particular vibrational coordinate (in the Herzberg-Teller spirit). Here we demonstrate that this effect is responsible for the 2PA spectral shapes of many fluorescent dyes including rhodamines, ATTO dyes, fluorescein, oxazines, acridines, pyronines, resorufin, and thionine. By measuring the two-photon polarization ratio as a function of excitation wavelength in these dyes, we were able to separate the transitions with Deltamu perpendicular to the long molecular axis (“FC”-type) and parallel to it (“HT”-type). We have found the previously undetected “HT-ghost” transitions under the pure electronic envelope. These are the 0-0 transitions of the "HT" progression build on the BLA frequency that are non-vanishing if the corresponding Huang-Rhys factor is not zero.

Quantum light allows imaging with sensitivity, speed and resolution beyond the reach of other techniques. Applications in biology are particularly important, because the best conventional microscopes are often severely constrained by photodamage to the specimen. Here, I will present experiment work from my lab applying squeezed light, a form of quantum light, to imaging of molecular vibrations. If time permits, I will also discuss work on quantum single-molecule imaging.

The rate of two-photon absorption of time-frequency-entangled photon pairs has been the subject of much study for its potential to enable quantum-enhanced molecular spectroscopy and imaging. We closely replicated recent experiments that reportedly observed such enhancement and have found that in the low-photon-flux regime the signal is below detection threshold. Using an optical parametric down-conversion photon-pair source that can be varied from the low-gain spontaneous regime to the high-gain squeezing regime, we observe two-photon absorption with a molecular sample in solution for the high-gain regime but not for the low-gain regime. The observed rates are consistent with theoretical predictions and indicate that time-frequency photon entanglement does not yet provide a practical means to enhance spectroscopy or imaging with current techniques.

We discuss advances in ultrasensitive Raman-spectroscopic probing of various biosamples. Our approach is based on laser spectroscopy aided by plasmonic nanoantennas, as for example in tip-enhanced Raman spectroscopy. An additional enhancement in sensitivity and speed is obtained by employing quantum molecular coherence driven either by pump and Stokes pulse pair or a direct infrared drive field.

Next generation single-photon detectors are becoming available. Research grade SPAD arrays promise camera-like detectors with dozens to hundreds of thousands of pixels, providing both spatial and temporal information about single photon events. Concurrently, other detector technologies are under development, such as crossed-delay line detectors. We evaluate the application of SPAD arrays and NCam, a crossed-delay line detector, towards quantum ghost imaging applications. We discuss implementation challenges and performance differences in the context of ghost imaging.

Modal imaging can provide unprecedented resolution for imaging point sources, such as single-molecule fluorescent tags used to study biological samples. Among different modal imaging techniques, image inversion interferometry, where the input field is separated into symmetric and antisymmetric components, in principle allows for resolving two point sources near the quantum limit. We investigate the implementation of inversion interferometry for superresolving point sources in fluorescence microscopy. Our work focuses on the integration of this technique with microscopes used in superresolution microscopy for imaging broadband point sources and fluorophores commonly used to tag proteins in cells.

We present progress towards demonstrating multi-passed stimulated Raman scattering microscopy and multi-passed transmission electron microscopy. A multi-pass microscope interrogates a sample multiple times in a cyclical and deterministic fashion. This can lead to a metrological advantage for imaging weak scatterers. The enhanced sensitivity can yield a significant reduction in damage imparted to biological samples or can reduce image acquisition time. The approach compares favorably with imaging techniques using squeezed or entangled probe states, but avoids the technical complexity associated with the production of such states.

Photosynthetic light harvesting in vivo displays near unit quantum efficiency under ultra-weak illumination conditions. A new generation of experimental and theoretical studies using quantum light sources and coincidence counting now allows explicit study of the absorption of individual photons. First generation experiments reveal a cycle of single photon absorption and single photon fluorescent emission that validates the microscopic interpretation of conventional bulk measurements of quantum efficiency. I shall discuss how these techniques and related theoretical studies can probe the spatiotemporal dynamics of photosynthesis in a new and fundamental manner.

We developed an ultrabroadband 2D electronic spectrometer that can measure ultrafast photophysics across the visible range. We apply this technique to explore the dissipative photophysical processes in the major light-harvesting protein in green plants, a mechanism by which plants protect against photooxidative damage.

Using a photon-counting quantum light spectroscopy that probes photosynthetic light harvesting with a single photon at a time, we experimentally demonstrated that photosynthesis begins and proceeds with a single quantum of energy. We report the observation of individual single-photon absorption and emission events in spatially distinct regions in photosynthetic systems. The experiments were carried out on an ensemble of pigment-protein complexes light-harvesting 2 from purple bacteria Rhodobacter sphaeroides under ambient conditions in vitro.

Quantum diamond microscopy (QDM) is a technique for wide field-of-view diffraction-limited imaging of a thin and dense layer of negatively charged nitrogen-vacancy (NV) defects in engineered diamond crystals. In the recent past, QDM has enabled imaging of a diverse class of samples with microscopic magnetic field profiles, for example, biological cells with magnetic nanoparticles and current flow profiles in thin quantum materials. A challenging extension of the technique is to perform spatially resolved magnetic field imaging from in-vitro 2D or 3D cultures of mammalian neurons. This potential development can lead to an alternative magnetometry-based neuronal imaging technique, with advantages over conventional scattering-limited multi-photon in-vivo calcium imaging.In this talk I will describe our path toward volume imaging of neurons with QDM.

The development of fluorescent molecular sensors for imaging voltage changes in biological systems has revolutionized neuroscience, providing a tool to capture neuronal activity over large areas with sub-neuron resolution both in vitro and in vivo [1–3]. However, the poor photostability of molecular voltage sensors limits recording times to a few minutes [1–3], posing problems for longitudinal studies of network evolution and disease processes. Here, we present an alternate non-invasive platform for sensitive high resolution voltage imaging using fluorescent, charge-sensitive defects in a transparent diamond substrate [6]. Using these charge state sensors, we establish an all-optical diamond voltage imaging microscope (DVIM) capable of sub-millisecond voltage imaging with sub-millivolt sensitivity. Our work open new pathways for the study of 2D neuronal network cultures as well as 3D brain organoids.

Quantum sensing tools have emerged as a compelling means to study nanoscale chemical and biological processes with high sensitivity and spatial resolution, promising wide impact in a variety of fields ranging from chemical synthesis to bioengineering. Recently there has been expanded interest in assay-like quantum sensing approaches that can yield high-fidelity analyte discrimination for practical applications. In this work, we introduce a novel high-throughput, in-flow, quantum sensing platform based on droplet microfluidics. Quantum sensors based on nanodiamonds hosting Nitrogen Vacancy (NV) centers are incorporated within monodisperse phase separated droplets which serve as picoliter containers for both the sensors and for analytes of interest. Such controllable micro-compartments allow for strong sensor-analyte interaction, and allow for the rapid, high throughput (~10kHz), quantum sensing of numerous droplets. We demonstrate a novel method for noise-suppression in the fluorescence-based optically detected magnetic resonance (ODMR) measurements exploiting droplet flow, and use it to carry out high sensitivity assay detection of several analytes of biological importance.

Iron is an essential yet toxic redox active element that is found in many cells, including neurons and glial cells. Several techniques have been used to quantify iron in neurons and cells; however, most are incapable of high-resolution imaging inside a single cell. Magnetic field sensors based on diamond nitrogen vacancy (NV) centers have emerged as a powerful tool for detecting magnetic signal in iron-containing biological samples with a good combination of spatial resolution and sensitivity. In this study we use NV based T1 relaxometry technique to map iron in cytochrome C (Cyt C) proteins. Cyt C plays an important role in the electron transport chain of mitochondria and it is in the Fe(III) paramagnetic state under ambient conditions. We measure Cyt C under different concentrations and locations of the 10-nm NV doped diamond chip. We show a reduction of the NV T1 from few milliseconds to hundreds of microseconds, explained by the spin noise from the intracellular iron spins in the Cyt C protein. Additionally, we perform imaging of Cyt C proteins on a nanostructured diamond chip.

We demonstrated single-photon emission from hBN color centers embedded inside live cells and their application to cellular barcoding. Each color center can exist in one out of 470 possible distinguishable states. A combination of a few color centers per cell can be used to uniquely tag virtually an unlimited number of cells. This barcoding technique is superior to others in almost all respects, including ease of production by a single-step procedure, biocompatibility and biodegradability, emission stability, no photobleaching, small size and a huge number of unique barcodes.

Imagine driving cell activities to treat injuries and disease simply by using tailored magnetic fields. Many relevant physiological processes, such as: the regulation of oxidative stress, proliferation, and respiration rates in cells; wound healing; ion channel functioning; and DNA repair were all demonstrated to be controlled by weak magnetic fields (with a strength on the order of that produced by your cell phone). Such macroscopic physiological responses to magnetic fields are consistent with being driven by chemical reactions that depend on the electron quantum property of spin. In the long-term, the electromagnetic fine-tuning of endogenous “quantum knobs” existing in nature could enable the development of drugs and therapeutic devices that could heal the human body — in a way that is non-invasive, remotely actuated, and easily accessible by anyone with a mobile phone. However, whereas spin-dependent chemical reactions have been unambiguously established for test-tube chemistry (bearing uncanny similarities with what physicists call “spin quantum sensing”), current research has not been able to deterministically link spin states to physiological outcomes in vivo and in real time. With novel quantum instrumentation, we are learning to control spin states within cells and tissues, having as a goal to write the “codebook” on how to deterministically alter physiology with weak magnetic fields to therapeutic and technological advantage.

In this talk, I will introduce metasurface-enhanced multiplexed nanospectroscopy and molecular diagnostics. First, we report metasurfaces-driven hyperspectral imaging via multiplexed plasmon resonance energy transfer to probe biological light-matter interactions, which can detect quantum biological electron transfer. We conduct real-time and label-free monitoring of reactive oxygen species (ROS) from different types of cells. We used three different cells, normal cells, tumor cells, and drug-treated tumor cells, and quantitatively analyzed ROS production in real-time. Second, we scalably fabricate a low-power-consuming metaphotonic PCR device composed of a metamaterial perfect absorber that can rapidly go through thermocycling steps using a single infrared LED. We demonstrate an ultrafast photonic reverse transcription PCR (RT-PCR) with 30 thermocycles achieving 21.9 °Cs-1 heating rate and 7.8 °Cs-1 cooling rate. Furthermore, we would like to utilize the metaphotonic PCR chip for quantitative quantum enzymology.

Confocal fluorescence microscopy is a common imaging technique for biomedical applications such as tumor characterization, precision resections, etc... However, the imaging process is limited by photo bleaching of samples and optical scattering limits imposed by wavelength limitations of traditionally used detection technologies. Here we show that by leveraging the high detection efficiency and large spectral range of superconducting nanowire single-photon detectors, it is possible to image deeper into tissue with a significant reduction in laser excitation power compared to the same technique using silicon photo-multipliers.

We've discovered a simple, nontoxic, biocompatible way to control the brightness of GFP-like fluorescent proteins via modest magnetic fields (~10 mT). Fluorescent proteins which seem magnetically inert (e.g. EGFP, mScarlet) become magnetoresponsive in the presence of an appropriate cofactor (e.g. EGFP-FlavinTag, or an mScarlet/FMN solution). This method works at room-temperature and body-temperature, in vitro, in E. coli and in cultured mammalian cells. The GFP-family magnetoresponse is weak (ΔF/F≈1%), but shows the hallmarks of evolvability. This suggests exciting technological possibilities, both short-term (e.g. lock-in detection, multiplexing) and long-term (e.g. optically-detected MRI, magnetogenetics). We've also discovered weak magnetoresponse from a member of the LOV-domain family. This suggests the possibility that magnetoresponse is a general feature of fluorescent proteins, and not unique to the cryptochrome/photolyase family.

The study of decoherence in biological systems has become an essential area of research for quantum mechanics. Fluorescence quenching is a physiochemical process that causes the decrease in fluorescence intensity of a fluorescent sample. Changing the surrounding environment of fluorescent proteins (FPs), by adding a fluorescence quenching agent could potentially have a large impact on the dynamical processes within these biological systems. Environmental influence on protein-protein interactions is not a well understood process, when looking at energy transfer dynamics of FPs. we attempt to establish the impact on the energy transfer dynamics between fluorophores by introducing varying concentrations of external quenchers to the FPs environment. We examine the photophysical properties and spectral changes of the FPs, after adding external quenchers to both the monomeric and dimeric structures to further characterise the energy transfer dynamics.

Quantum mechanics has significantly advanced our understanding of fundamental properties. While biological studies have traditionally been assumed to be governed by classical physics, recent studies have shown that coherent excitonic coupling between two chromophores in a homodimer of the yellow fluorescent protein, Venus, is possible at room temperature [Y. Kim et al., Biophys. J., 2019, 116, 1918-1930]. In this study, we present a genetically encoded sensor, inspired by recent finding of the ultrafast photoinduced energy transfer between dimeric enhanced green fluorescent proteins (dEGFPs), to monitor hydrophobic environmental changes [A. Sanchez-Pedreno Jimenez, et al., Phys. Chem. Chem. Phys., in press]. This sensor can be used to investigate the spatial and temporal dynamics biocondensates in cells. This biosensor offers an innovative approach to unravel the dynamics of biocondensates, to elucidate their biological functions and potential implications in health and disease. Experiments are being conducted to test this sensor in living cells.