All forms of microscopy that have previously been developed rely on classical sources of light, which are relatively noisy by virtue of the photons arriving randomly in time and position. We propose a new technique of two-beam optical microscopy using a nonclassical light source comprising entangled photon pairs generated by the process of nonlinear optical parametric downconversion. Entangled-photon microscopy (EPM) has the potential for providing a substantial improvement over confocal laser scanning fluorescence microscopy (CLSM) as well as over two-photon laser scanning fluorescence microscopy (TPLSM). The latter technique relies on the proportionality of the two-photon absorption rate to the square of the optical photon-flux density, and makes use of a highly focused optical beam to localize the region from which fluorescence is observed. However, because the arrival of pairs of photons is accidental in TPLSM, relatively large values of the photon-flux density are required to achieve photon coincidence and hence absorption. This can lead to undesired phototoxicity and photobleaching of the specimen. EPM, in contrast, relies on the presence of highly correlated photon pairs, which enhances the two-photon absorption rate significantly and enables lower values of the photon-flux density to be used. The process depends linearly, rather than quadratically, on the photon-flux density. Further, the EPM interaction region can be controlled by the use of two beams, providing increased flexibility in the selection of desired regions of a specimen, and leading to enhanced axial and lateral resolution.
All forms of microscopy that have previously been developed rely on classical sources of light, which are relatively noisy by virtue of the photons arriving randomly in time and position. We propose a new technique of two-beam optical microscopy using a nonclassical light source comprising entangled photon pairs generated by the process of nonlinear optical parametric downconversion. Entangled-photon microscopy (EPM) has the potential for providing a substantial improvement over confocal laser scanning fluorescence microscopy (CLSM) as well as over two-photon laser scanning fluorescence microscopy (TPLSM). The latter technique relies on the proportionality of the two-photon absorption rate to the square of the optical photon-flux density, and makes use of a highly focused optical beam to localize the region from which fluorescence is observed. However, because the arrival of pairs of photons is accidental in TPLSM, relatively large values of the photon-flux density are required to achieve photon coincidence and hence absorption. This can lead to undesired phototoxicity and photobleaching of the specimen. EPM, in contrast, relies on the presence of highly correlated photon pairs, which enhances the two-photon absorption rate significantly and enables lower values of the photon-flux density to be used. The process depends linearly, rather than quadratically, on the photon-flux density. Further, the EPM interaction region can be controlled by the use of two beams, providing increased flexibility in the selection of desired regions of a specimen, and leading to enhanced axial and lateral resolution.
Fluorescence microscopy has become integral to biological studies for the technique’s ability to elucidate structures of biomolecules for in-situ studies with high selectivity and specificity. Imaging of intrinsic indicators, such as fluorescent amino acids in proteins, provides important information, but can be challenging to accomplish. Current microscopy techniques that measure native fluorescence without the use of exogenous labels involve either direct UV excitation which is commonly non-localized and can be detrimental to the system, or multiphoton absorption which must be conducted at high intensities, therefore posing high risks of photodamage. As such, we seek to investigate an efficient way to gently excite native fluorescence in biological systems in a way that overcomes these limitations. Quantum entangled photon pairs generated via spontaneous parametric downconversion (SPDC), may be an alternative to conducting two-photon absorption (TPA) to excite fluorescence in amino acids without the high fluences currently used. These photon pairs are highly correlated in time. Thus, the arrival of one photon is simultaneously followed by the arrival of its sister photon. As a result, a molecule interacting with the photon pair should simultaneously absorb both photons, leading to a linear two-photon absorption rate, and the linearity of the two-photon process should dramatically reduce the light intensity necessary for TPA. Therefore, quantum entangled photon pairs offer the possibility of performing low intensity UV excitation using photons in the visible wavelength range. With this work, we generated and characterized entangled photons generated via SPDC, and investigated whether fluorescent amino acids can be excited, and the subsequent fluorescence induced with entangled two-photon absorption. Results show that much higher entangled two-photon rates than what are currently available are needed to measure significant signals with entangled two-photon excitation.
Nonlinear optics has had extensive application into a vast array of scientific fields. One such nonlinear process, two-photon absorption (TPA), has had a wildly successful \nadoption into the field of biological imaging and microscopy. As far and as fast as this field is progressing, limitations stemming from the use of ultrafast lasers are starting to appear. \nIn this work, an alternative nonclassical light source will be motivated for the application \nof low photon-flux two-photon microscopy. The origin and properties of the chosen \nnonclassical source, spontaneous parametric downconversion (SPDC), will be discussed along with the spatial and spectral properties modelled. Nonlinear processes such as TPA and sum frequency generation (SFG) will be viewed as "ultrafast coincidence measurements" of two photons arriving at a molecule within the time window of excitation. These ultrafast coincidence measurements will be viewed in an alternative manner: in terms of the second-order coherence from a light source. This degree of second-order coherence can be subdivided into two categories arising from different combinations of correlations within and between entangled photon pairs. Of interest, the energy-time correlations within the photon pair allow for enhancements in ultrafast coincidence rates over coherent light sources. The makings of an experimental setup to demonstrate enhanced rates from ultrafast two-photon coincidences taking place in SFG in a nonlinear crystal will be discussed.
Quantum entanglement has been shown to imply correlations stronger than those allowed by classical models. The possibility of performing tasks that are classically impossible has made quantum entanglement a powerful resource for the development of novel methods and applications in various fields of research such as quantum computing, quantum cryptography, and quantum metrology. There is a great need for the development of next generation instrumentation and technologies utilizing entangled quantum light. Among the many applications of nonclassical states of light, nonlinear microscopy has the potential to make an impact in broad areas of science from physics to biology. Here, the microscopic image created by the fluorescence selectively excited by the process of the entangled two-photon absorption is reported. Entangled two-photon microscopy offers nonlinear imaging capabilities at an unprecedented low excitation intensity 10 7, which is 6 orders of magnitude lower than the excitation level for the classical two-photon image. The nonmonotonic dependence of the image on the femtosecond delay between the components of the entangled photon pair is demonstrated. This delay dependence is a result of specific quantum interference effects associated with the entanglement and this is not observable with classical excitation light. In combination with novel spectroscopic capabilities provided by a nonclassical light excitation, this is of critical importance for sensing and biological applications.
Correct biological interpretation from cell imaging can be achieved only if the observed phenomena proceed with negligible perturbation from the imaging system. Herein, we demonstrate microscopic images of breast cancer cells created by the fluorescence selectively excited in the process of entangled two-photon absorption in a scanning microscope at an excitation intensity orders of magnitude lower than that used for classical two-photon microscopy. Quantum enhanced entangled two-photon microscopy has shown cell imaging capabilities at an unprecedented low excitation intensity of ∼3.6 × 10 7 photons/s, which is a million times lower than the excitation level for the classical two-photon fluorescence image obtained in the same microscope. The extremely low light probe intensity demonstrated in entangled two-photon microscopy is of critical importance to minimize photobleaching during repetitive imaging and damage to cells in live-cell applications. This technology opens new avenues in cell investigations with light microscopy, such as enhanced selectivity and time–frequency resolution.
The harnessing of the nonclassical properties in photon pairs generated by the process of Spontaneous Parametric Down Conversion (SPDC) to enhance two-photon interactions has long intrigued researchers. Their use in light-matter interactions has been proven to improve efficiency either by increasing the rate of generation or by suppressing noise. Namely, the processes of Entangled Sum Frequency Generation (eSFG) and Entangled Two Photon Absorption (eTPA) have been shown to produce more photons when stimulated with entangled photons than the classical versions of the effects. This has only been effective so far at very low intensities, which, alongside the inability to reproduce previous experimental results, has caused debates about the usefulness of the effect. In the presented work, I aim to shed light on the topic by employing a carefully crafted source of entangled photon pairs to improve two-photon effects. I attempt to demonstrate, through a series of experiments, that the quantum enhancement can be observed at an order of magnitude higher than previously thought possible. To solidify the results, a direct comparison of the quantum effect was conducted by investigating the performance of the phenomenon when it is stimulated by classical radiation from a laser. Our results confirm a quantum improvement in a regime previously considered impossible, and they enabled us to examine the ramifications of the effect on imaging. By doing so, we consider real-life microscopy applications, and therefore, we can set limits to the effect.
The application of quantum states of light such as entangled photons, for example, created by parametric down conversion, has experienced tremendous progress in the almost 40 years since their first experimental realization. Initially, they were employed in the investigation of the foundations of quantum physics, such as the violation of Bell's inequalities and studies of quantum entanglement. They later emerged as basic platforms for quantum communication protocols and, in the recent experiments on single-photon interactions, in photonic quantum computation. These applications aim at the controlled manipulation of the photonic degrees of freedom, and therefore rely on simple models of matter, where the analysis is simpler. Furthermore, quantum imaging with entangled light can achieve enhanced resolution, and quantum metrology can overcome the shot noise limit for classical light. This Account focuses on an entirely different emerging class of applications using quantum light as a powerful spectroscopic tool to reveal novel information about complex molecules. These applications utilize two appealing properties of quantum light: its distinct intensity fluctuations and its nonclassical bandwidth properties. These give rise to new and surprising behavior of nonlinear optical signals. Nonclassical intensity fluctuations can enhance nonlinear optical signals relative to linear absorption. For instance, the two-photon absorption of entangled photon pairs scales linearly (rather than quadratically) in the photon flux, just like a single photon absorption. This enables nonlinear quantum spectroscopy of photosensitive, for example, biological, samples at low light intensities. We will discuss how the two-photon absorption cross section becomes a function of the photonic quantum state, which can be manipulated by properties of the entangled photon pairs. In addition, the quantum correlations in entangled photon states further influence the nonlinear signals in a variety of ways. Apart from affecting the signal's scaling with intensity, they also constitute an entirely new approach to shaping and controlling excitation pathways in molecular aggregates in a way that cannot be achieved with shaped classical pulses. This is because between the two absorption events in entangled two-photon absorption, the light and material system are entangled. Classical constraints for the simultaneous time and frequency resolution can thus be circumvented, since the two are not Fourier conjugates. Here we review the simplest manifestation of quantum light spectroscopy, two-photon absorption spectroscopy with entangled photons. This will allow us to discuss exemplarily the impact of quantum properties of light on a nonlinear optical signal and explore the opportunities for future applications.