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Quantitative phase microscopies (QPMs) play a pivotal role in bio-imaging, offering unique insights that complement fluorescence imaging. They provide essential data on mass distribution and transport, inaccessible to fluorescence techniques. Additionally, QPMs are label-free, eliminating concerns of photobleaching and phototoxicity. However, navigating through the array of available QPM techniques can be complex, making it challenging to select the most suitable one for a particular application. This tutorial review presents a thorough comparison of the main QPM techniques, focusing on their accuracy in terms of measurement precision and trueness. We focus on 8 techniques, namely digital holographic microscopy (DHM), cross-grating wavefront microscopy (CGM), which is based on QLSI (quadriwave lateral shearing interferometry), diffraction phase microscopy (DPM), differential phase-contrast (DPC) microscopy, phase-shifting interferometry (PSI) imaging, Fourier phase microscopy (FPM), spatial light interference microscopy (SLIM), and transport-of-intensity equation (TIE) imaging. For this purpose, we used a home-made numerical toolbox based on discrete dipole approximation (IF-DDA). This toolbox is designed to compute the electromagnetic field at the sample plane of a microscope, irrespective of the object's complexity or the illumination conditions. We upgraded this toolbox to enable it to model any type of QPM, and to take into account shot noise. In a nutshell, the results show that DHM and PSI are inherently free from artefacts and rather suffer from coherent noise; In CGM, DPC, DPM and TIE, there is a trade-off between precision and trueness, which can be balanced by varying one experimental parameter; FPM and SLIM suffer from inherent artefacts that cannot be discarded experimentally in most cases, making the techniques not quantitative especially for large objects covering a large part of the field of view, such as eukaryotic cells., (© 2024. The Author(s).)

The article introduces an optical microscopy technique capable of simultaneously acquiring quantitative fluorescence and phase (or equivalently wavefront) images with a single camera sensor, avoiding any delay between both images, or registration of images acquired separately. The method is based on the use of a 2-dimensional diffraction grating (aka cross-grating) positioned at a millimeter distance from a 2-color camera. Fluorescence and wavefront images are extracted from the two color channels of the camera, and retrieved by image demodulation. The applicability of the method is illustrated on various samples, namely fluorescent micro-beads, bacteria and mammalian cells., (© 2024. The Author(s).)

Gold nanoparticles (AuNPs) are increasingly used for their thermoplasmonic properties, i.e., their ability to convert light energy into heat through plasmon resonance. However, measuring temperature gradients generated at the microscale by assemblies of AuNPs remains challenging, especially for random 3D distributions of AuNPs. Here, we introduce a label-free thermometry approach, combining quantitative wavefront microscopy and numerical simulations, to infer the heating power dissipated by a 3D model system consisting of emulsion microdroplets loaded with AuNPs. This approach gives access to the temperature reached in the droplets under laser irradiation without the need for extrinsic calibration. This versatile thermometry method is promising for noninvasive temperature measurements in various 3D microsystems involving AuNPs as colloidal heat sources, including photothermal drug delivery systems.

Quantitative phase microscopy (QPM) represents a noninvasive alternative to fluorescence microscopy for cell observation with high contrast and for the quantitative measurement of dry mass (DM) and growth rate at the single-cell level. While DM measurements using QPM have been widely conducted on mammalian cells, bacteria have been less investigated, presumably due to the high resolution and high sensitivity required by their smaller size. This article demonstrates the use of cross-grating wavefront microscopy, a high-resolution and high-sensitivity QPM, for accurate DM measurement and monitoring of single microorganisms (bacteria and archaea). The article covers strategies for overcoming light diffraction and sample focusing, and introduces the concepts of normalized optical volume and optical polarizability (OP) to gain additional information beyond DM. The algorithms for DM, optical volume, and OP measurements are illustrated through two case studies: monitoring DM evolution in a microscale colony-forming unit as a function of temperature, and using OP as a potential species-specific signature., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 Biophysical Society. Published by Elsevier Inc. All rights reserved.)

Quantitative phase microscopies (QPMs) enable label-free, non-invasive observation of living cells in culture, for arbitrarily long periods of time. One of the main benefits of QPMs compared with fluorescence microscopy is the possibility to measure the dry mass of individual cells or organelles. While QPM dry mass measurements on neural cells have been reported this last decade, dry mass measurements on their neurites has been very little addressed. Because neurites are tenuous objects, they are difficult to precisely characterize and segment using most QPMs. In this article, we use cross-grating wavefront microscopy (CGM), a high-resolution wavefront imaging technique, to measure the dry mass of individual neurites of primary neurons in vitro . CGM is based on the simple association of a cross-grating positioned in front of a camera, and can detect wavefront distortions smaller than a hydrogen atom (∼0.1 nm). In this article, an algorithm for dry-mass measurement of neurites from CGM images is detailed and provided. With objects as small as neurites, we highlight the importance of dealing with the diffraction rings for proper image segmentation and accurate biomass measurements. The high precision of the measurements we obtain using CGM and this semi-manual algorithm enabled us to detect periodic oscillations of neurites never observed before, demonstrating the sufficient degree of accuracy of CGM to capture the cell dynamics at the single neurite level, with a typical precision of 2%, i.e., 0.08 pg in most cases, down to a few fg for the smallest objects., Competing Interests: The authors declare no conflicts of interest., (© 2022 Optica Publishing Group under the terms of the Optica Open Access Publishing Agreement.)

Thermophiles are microorganisms that thrive at high temperature. Studying them can provide valuable information on how life has adapted to extreme conditions. However, high temperature conditions are difficult to achieve on conventional optical microscopes. Some home-made solutions have been proposed, all based on local resistive electric heating, but no simple commercial solution exists. In this article, we introduce the concept of microscale laser heating over the field of view of a microscope to achieve high temperature for the study of thermophiles, while maintaining the user environment in soft conditions. Microscale heating with moderate laser intensities is achieved using a substrate covered with gold nanoparticles, as biocompatible, efficient light absorbers. The influences of possible microscale fluid convection, cell confinement and centrifugal thermophoretic motion are discussed. The method is demonstrated with two species: (i) Geobacillus stearothermophilus, a motile thermophilic bacterium thriving around 65 °C, which we observed to germinate, grow and swim upon microscale heating and (ii) Sulfolobus shibatae, a hyperthermophilic archaeon living at the optimal temperature of 80 °C. This work opens the path toward simple and safe observation of thermophilic microorganisms using current and accessible microscopy tools., (© 2022. The Author(s).)

Optically-assisted large-scale assembly of nanoparticles have been of recent interest owing to their potential in applications to assemble and manipulate colloidal particles and biological entities. In the recent years, plasmonic heating has been the most popular mechanism to achieve temperature hotspots needed for extended assembly and aggregation. In this work, we present an alternative route to achieving strong thermal gradients that can lead to non-equilibrium transport and assembly of matter. We utilize the excellent photothermal properties of graphene oxide to form a large-scale assembly of silica beads. The formation of the assembly using this scheme is rapid and reversible. Our experiments show that it is possible to aggregate silica beads (average size 385 nm) by illuminating thin graphene oxide microplatelet by a 785 nm laser at low intensities of the order of 50-100 µW/µm 2 . We further extend the study to trapping and photoablation of E. coli bacteria using graphene oxide. We attribute this aggregation process to optically driven thermophoretic forces. This scheme of large-scale assembly is promising for the study of assembly of matter under non-equilibrium processes, rapid concentration tool for spectroscopic studies such as surface-enhanced Raman scattering and for biological applications., (© 2022. The Author(s).)

Whereas heating nanoparticles with light is straightforward, measuring the resulting nanoscale temperature increase is intricate and still a matter of active research in plasmonics, with envisioned applications in nanochemistry, biomedicine, and solar light harvesting, among others. Interestingly, this research line mostly belongs to the optics community today because light is not only used for heating but also often for probing temperature. In this Perspective, I present and discuss recent advances in the search for efficient and reliable thermometry techniques for nanoplasmonic systems by the nano-optics community. I focus on the recently proposed approach based on the spectral measurement of anti-Stokes emission from the plasmonic nanoparticles themselves.

Culturing cells confined in microscale geometries has been reported in many studies this last decade, in particular following the development of microfluidic-based applications and lab-on-a-chip devices. Such studies usually examine growth of Escherichia coli. In this article, we show that E. coli may be a poor model and that spatial confinement can severely prevent the growth of many micro-organisms. By studying different bacteria and confinement geometries, we determine that the growth inhibition observed for some bacteria results from fast dioxygen depletion, inherent to spatial confinement, and not to any depletion of nutriments. This article unravels the physical origin of confinement problems in cell culture, highlighting the importance of oxygen depletion, and paves the way for the effective culturing of bacteria in confined geometries by demonstrating enhanced cell growth in confined geometries in the proximity of air bubbles., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Plasmonic nanotweezers use intense electric field gradients to generate optical forces able to trap nano-objects in liquids. However, part of the incident light is absorbed into the metal, and a supplementary thermophoretic force acting on the nano-object arises from the resulting temperature gradient. Plasmonic nanotweezers thus face the challenge of disentangling the intricate contributions of the optical and thermophoretic forces. Here, we show that commonly added surfactants can unexpectedly impact the trap performance by acting on the thermophilic or thermophobic response of the nano-object. Using different surfactants in double nanohole plasmonic trapping experiments, we measure and compare the contributions of the thermophoretic and the optical forces, evidencing a trap stiffness 20× higher using sodium dodecyl sulfate (SDS) as compared to Triton X-100. This work uncovers an important mechanism in plasmonic nanotweezers and provides guidelines to control and optimize the trap performance for different plasmonic designs.

Over the past two decades, there has been a growing interest in the use of plasmonic nanoparticles as sources of heat remotely controlled by light, giving rise to the field of thermoplasmonics. The ability to release heat on the nanoscale has already impacted a broad range of research activities, from biomedicine to imaging and catalysis. Thermoplasmonics is now entering an important phase: some applications have engaged in an industrial stage, while others, originally full of promise, experience some difficulty in reaching their potential. Meanwhile, innovative fundamental areas of research are being developed. In this Review, we scrutinize the current research landscape in thermoplasmonics, with a specific focus on its applications and main challenges in many different fields of science, including nanomedicine, cell biology, photothermal and hot-electron chemistry, solar light harvesting, soft matter and nanofluidics.

Light absorption and scattering of plasmonic metal nanoparticles can lead to non-equilibrium charge carriers, intense electromagnetic near-fields, and heat generation, with promising applications in a vast range of fields, from chemical and physical sensing to nanomedicine and photocatalysis for the sustainable production of fuels and chemicals. Disentangling the relative contribution of thermal and non-thermal contributions in plasmon-driven processes is, however, difficult. Nanoscale temperature measurements are technically challenging, and macroscale experiments are often characterized by collective heating effects, which tend to make the actual temperature increase unpredictable. This work is intended to help the reader experimentally detect and quantify photothermal effects in plasmon-driven chemical reactions, to discriminate their contribution from that due to photochemical processes and to cast a critical eye on the current literature. To this aim, we review, and in some cases propose, seven simple experimental procedures that do not require the use of complex or expensive thermal microscopy techniques. These proposed procedures are adaptable to a wide range of experiments and fields of research where photothermal effects need to be assessed, such as plasmonic-assisted chemistry, heterogeneous catalysis, photovoltaics, biosensing, and enhanced molecular spectroscopy., Competing Interests: Conflict of interestThe authors declare that they have no conflict of interest., (© The Author(s) 2020.)

Gold films do not adhere well on glass substrates, so plasmonics experiments typically use a thin adhesion layer of titanium or chromium to ensure a proper adhesion between the gold film and the glass substrate. While the absorption of light into gold structures is largely used to generate heat and control the temperature at the nanoscale, the influence of the adhesion layer on this process is largely overlooked. Here, we quantify the role of the adhesion layer in determining the local temperature increase around a single nanohole illuminated by a focused infrared laser. Despite their nanometer thickness, adhesion layers can absorb a greater fraction of the incoming infrared light than the 100 nm thick gold layer leading to a significant increase of the local temperature. Different experimental designs are explored, offering new ways to promote or avoid the temperature increase inside nanoapertures. This knowledge further expands the plasmonic toolbox for temperature-controlled experiments including single molecule sensing, nanopore translocation, polymerization, or nano-optical trapping.

In this paper, we introduce a formalism to determine the relationship between the full vectorial electric field existing at the object plane of a microscope and that existing at the image plane. The model is then used to quantitatively simulate, in both phase and intensity, the image of a radiating electric dipole placed either in a homogeneous medium or in the vicinity of a substrate. These simulations are compared with experimental measurements on single gold nanoparticles carried out by quadriwave lateral shearing interferometry.

Heating on the microscale using focused lasers gave rise to recent applications, e.g., in biomedicine, biology and microfluidics, especially using gold nanoparticles as efficient nanoabsorbers of light. However, such an approach naturally leads to nonuniform, Gaussian-like temperature distributions due to the diffusive nature of heat. Here, we report on an experimental means to generate arbitrary distributions of temperature profiles on the micrometric scale (e.g. uniform, linear, parabolic, etc) consisting in illuminating a uniform gold nanoparticle distribution on a planar substrate using spatially contrasted laser beams, shaped using a spatial light modulator (SLM). We explain how to compute the light pattern and the SLM interferogram to achieve the desired temperature distribution, and demonstrate the approach by carrying out temperature measurements using quantitative wavefront sensing.

Laser heating of individual cells in culture recently led to seminal studies in cell poration, fusion, migration, or nanosurgery, although measuring the local temperature increase in such experiments remains a challenge. Here, the laser-induced dynamical control of the heat-shock response is demonstrated at the single cell level, enabled by the use of light-absorbing gold nanoparticles as nanosources of heat and a temperature mapping technique based on quadriwave lateral shearing interferometry (QLSI) measurements. As it is label-free, this approach does not suffer from artifacts inherent to previously reported fluorescence-based temperature-mapping techniques and enables the use of any standard fluorescent labels to monitor in parallel the cell's response., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Metal nitrides have been proposed to replace noble metals in plasmonics for some specific applications. In particular, while titanium nitride (TiN) and zirconium nitride (ZrN) possess localized plasmon resonances very similar to gold in magnitude and wavelength, they benefit from a much higher sustainability to temperature. For this reason, they are foreseen as ideal candidates for applications in nanoplasmonics that require high material temperature under operation, such as heat assisted magnetic recording (HAMR) or thermophotovoltaics. This article presents a detailed investigation of the plasmonic properties of TiN and ZrN nanoparticles in comparison with gold nanoparticles, as a function of the nanoparticle morphology. As a main result, metal nitrides are shown to be poor near-field enhancers compared to gold, no matter the nanoparticle morphology and wavelength. The best efficiencies of metal nitrides as compared to gold in term of near-field enhancement are obtained for small and spherical nanoparticles, and they do not exceed 60%. Nanoparticle enlargements or asymmetries are detrimental. These results mitigate the utility of metal nitrides for high-temperature applications such as HAMR, despite their high temperature sustainability. Nevertheless, at resonance, metal nitrides behave as efficient nanosources of heat and could be relevant for applications in thermoplasmonics, where heat generation is not detrimental but desired.

Solvothermal synthesis, denoting chemical reactions occurring in metastable liquids above their boiling point, normally requires the use of a sealed autoclave under pressure to prevent the solvent from boiling. This work introduces an experimental approach that enables solvothermal synthesis at ambient pressure in an open reaction medium. The approach is based on the use of gold nanoparticles deposited on a glass substrate and acting as photothermal sources. To illustrate the approach, the selected hydrothermal reaction involves the formation of indium hydroxide microcrystals favored at 200 °C in liquid water. In addition to demonstrating the principle, the benefits and the specific characteristics of such an approach are investigated, in particular, the much faster reaction rate, the achievable spatial and time scales, the effect of microscale temperature gradients, the effect of the size of the heated area, and the effect of thermal-induced microscale fluid convection. This technique is general and could be used to spatially control the deposition of virtually any material for which a solvothermal synthesis exists., Competing Interests: The authors declare the following competing financial interest(s): HR has been a PhD student from April 2015 with financial support that came partly from the Phasics SA company.

In this article, we present a comprehensive investigation of the photothermal properties of plasmonic nanowire networks. We measure the local steady-state temperature increase, heat source density, and absorption in Ag, Au, and Ni metallic nanowire networks under optical illumination. This allows direct experimental confirmation of increased heat generation at the junction between two metallic nanowires and stacking-dependent absorption of polarized light. Due to thermal collective effects, the local temperature distribution in a network is shown to be completely delocalized on a micrometer scale, despite the nanoscale features in the heat source density. Comparison of the experimental temperature profile with numerical simulation allows an upper limit for the effective thermal conductivity of a Ag nanowire network to be established at 43 Wm(-1) K(-1) (0.1 κbulk).

We argue that standard thermodynamic considerations and scaling laws show that a single cell cannot substantially raise its temperature by endogenous thermogenesis. This statement seriously questions the interpretations of recent work reporting temperature heterogeneities measured in single living cells.

We introduce a deterministic procedure, named TSUNA for Temperature Shaping Using Nanoparticle Assemblies, aimed at generating arbitrary temperature distributions on the microscale. The strategy consists in (i) using an inversion algorithm to determine the exact heat source density necessary to create a desired temperature distribution and (ii) reproducing experimentally this calculated heat source density using smart assemblies of lithographic metal nanoparticles under illumination at their plasmonic resonance wavelength. The feasibility of this approach was demonstrated experimentally by thermal microscopy based on wavefront sensing.

Noble metal nanoparticles supporting plasmonic resonances behave as efficient nanosources of light, heat and energetic electrons. Owing to these properties, they offer a unique playground to trigger chemical reactions on the nanoscale. In this tutorial review, we discuss how nanoplasmonics can benefit chemistry and review the most recent developments in this new and fast growing field of research.

The fabrication of micro/nanostructured surfaces functionalized with stimulus-responsive chemical groups proved to be an interesting approach to simultaneously confine cell adhesion and manipulate cell-substrate interactions down to the single cell level. However, reversibility of stimulus-triggered systems is often not possible or exhibits slow switching kinetics. In contrast to such setups, gold nanoparticles have the properties to efficiently and reversibly generate heat under illumination at their plasmon resonance band. Thus, photo-induced heating could be used to directly and locally interface living cells and dynamically tailor the interactions to their adhesive environment. In the present chapter, we will first detail the preparation of micropatterned and functionalized gold nanoparticles immobilized on glass coverslips, and then report how to reliably characterize the photothermal properties of such substrates that enable the dynamic manipulation of cells., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The temperature distribution throughout arrays of illuminated metal nanoparticles is investigated numerically and experimentally. The two cases of continuous and femtosecond-pulsed illumination are addressed. In the case of continuous illumination, two distinct regimes are evidenced: a temperature confinement regime, where the temperature increase remains confined at the vicinity of each nanosource of heat, and a temperature delocalization regime, where the temperature is uniform throughout the whole nanoparticle assembly despite the heat sources' nanometric size. We show that the occurrence of one regime or another simply depends on the geometry of the nanoparticle distribution. In particular, we derived (i) simple expressions of dimensionless parameters aimed at predicting the degree of temperature confinement and (ii) analytical expressions aimed at estimating the actual temperature increase at the center of an assembly of nanoparticles under illumination, preventing heavy numerical simulations. All these theoretical results are supported by experimental measurements of the temperature distribution on regular arrays of gold nanoparticles under illumination. In the case of femtosecond-pulsed illumination, we explain the two conditions that must be fulfilled to observe a further enhanced temperature spatial confinement.

The ability to reversibly control the interactions between the extracellular matrix (ECM) and cell surface receptors such as integrins would allow one to investigate reciprocal signaling circuits between cells and their surrounding environment. Engineering microstructured culture substrates functionalized with switchable molecules remains the most adopted strategy to manipulate surface adhesive properties, although these systems exhibit limited reversibility and require sophisticated preparation procedures. Here, we report a straightforward protocol to fabricate biofunctionalized micropatterned gold nanoarrays that favor one-dimensional cell migration and function as plasmonic nanostoves to physically block and orient the formation of new adhesion sites. Being reversible and not restricted spatiotemporally, thermoplasmonic approaches will open new opportunities to further study the complex connections between ECM and cells.

Self-assembled plasmonic nanoparticle networks (PNN) composed of chains of 12 nm diameter crystalline gold nanoparticles exhibit a longitudinally coupled plasmon mode centered at 700 nm. We have exploited this longitudinal absorption band to efficiently confine light fields and concentrate heat sources in the close vicinity of these plasmonic chain networks. The mapping of the two phenomena on the same superstructures was performed by combining two-photon luminescence and fluorescence polarization anisotropy imaging techniques. Besides the light and heat concentration, we show experimentally that the planar spatial distribution of optical field intensity can be simply modulated by controlling the linear polarization of the incident optical excitation. On the contrary, the heat production, which is obtained here by exciting the structures within the optically transparent window of biological tissues, is evenly spread over the entire PNN. This contrasts with the usual case of localized heating in continuous nanowires, thus opening opportunities for these networks in light-induced hyperthermia applications. Furthermore, we propose a unified theoretical framework to account for both the nonlinear optical and thermal near-fields around PNN. The associated numerical simulations, based on a Green's function formalism, are in excellent agreement with the experimental images. This formalism therefore provides a versatile tool for the accurate engineering of optical and thermodynamical properties of complex plasmonic colloidal architectures.

Heat is of fundamental importance in many cellular processes such as cell metabolism, cell division and gene expression. (1-3) Accurate and noninvasive monitoring of temperature changes in individual cells could thus help clarify intricate cellular processes and develop new applications in biology and medicine. Here we report the use of green fluorescent proteins (GFP) as thermal nanoprobes suited for intracellular temperature mapping. Temperature probing is achieved by monitoring the fluorescence polarization anisotropy of GFP. The method is tested on GFP-transfected HeLa and U-87 MG cancer cell lines where we monitored the heat delivery by photothermal heating of gold nanorods surrounding the cells. A spatial resolution of 300 nm and a temperature accuracy of about 0.4 °C are achieved. Benefiting from its full compatibility with widely used GFP-transfected cells, this approach provides a noninvasive tool for fundamental and applied research in areas ranging from molecular biology to therapeutic and diagnostic studies., (© 2012 American Chemical Society)

We introduce an optical microscopy technique aimed at characterizing the heat generation arising from nanostructures, in a comprehensive and quantitative manner. Namely, the technique permits (i) mapping the temperature distribution around the source of heat, (ii) mapping the heat power density delivered by the source, and (iii) retrieving the absolute absorption cross section of light-absorbing structures. The technique is based on the measure of the thermal-induced refractive index variation of the medium surrounding the source of heat. The measurement is achieved using an association of a regular CCD camera along with a modified Hartmann diffraction grating. Such a simple association makes this technique straightforward to implement on any conventional microscope with its native broadband illumination conditions. We illustrate this technique on gold nanoparticles illuminated at their plasmonic resonance. The spatial resolution of this technique is diffraction limited, and temperature variations weaker than 1 K can be detected., (© 2012 American Chemical Society)