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Harmonic nanoparticles
The optical microscopy field is continuously evolving in order to improve the resolution and contrast of images. Numerous methods, such as confocal, dark-field, phase-contrast and polarization microscopy, are shown as improvements to conventional optical microscopy. As a visualization tool, ordinary optical microscopy suffers from its low physical and chemical specificity. This can be eliminated by combining microscopy with spectroscopic techniques such as fluorescence, infrared or Raman spectroscopy [5].
Nonlinear optical microscopy is based on the strong interaction of light and matter, revealing specific chemical / biological properties and structures. The contrast of the obtained images is determined by the difference in the nonlinear optical properties of the object and the environment.
Fig. 1.1 Energy diagram for linear (luminescence) and nonlinear optical processes: (a) photoluminescence, (b) two photon luminescence, (c) second harmonic generation, (d) third harmonic generation, (e) CARS – coherent anti-Stokes Raman scattering. Dashed lines corresponds to the virtual states [6].
The efficiency of the interaction of light with matter is characterized by the dielectric susceptibility of the material. When the objects are excited by the radiation of halogen lamps or continuous (CW) lasers, the electromagnetic response is linear in relation to the magnitude of the incident light field. However, with the use of pulsed laser radiation of high intensity, which usually exceeds several MW/cm2, a series of nonlinear optical effects, whose contributions depend on the intensity of excitation, are clearly manifested.
Typically, image formation in NLO microscopy is based on two types of NLO effects that result in the laser radiation frequency conversion: (i) the generation of optical harmonics, and (ii) the generation of combinational radiation through the excitation of vibrational resonances of chemical bonds. For the mentioned parametric processes, there is a generation of coherent radiation at wavelengths that differ from the wavelength of excitation [7].
Fig. 1.1 schematically presents energy diagrams for the following processes: photoluminescence with (a) single- and (b) two-photon excitation, generation of the second (SH) and third (TH) optical harmonics, and coherent anti-Stokes Raman scattering of light (CARS) [6]. Compared to fluorescence microscopy with single-photon excitation, multiphoton microscopy has an enhanced imaging depth and high spatial resolution (see Fig. 1.2) by reading the signal only from the region of the laser beam waist [8].
Fig. 1.2 Examples of simultaneously captured images using techniques of multiphoton microscopy for human breast cancer tissue excited in a transparency window: (a) two-photon fluorescence reflects fluorescence properties; (b) SHG – manifestation of noncentrosymmetry; (c) THG – manifestation of heterogeneity; (d) CARS is the chemical composition of the tissues [9].
In this context, Harmonic Nanoparticles (HNPs) are a new type of NLO markers for biological systems based on inorganic oxide nanocrystals with a noncentrosymmetric lattice that effectively convert laser radiation [10–12]. This term was introduced for the designation of a new broad class of nanoparticles (NPs) that can simultaneously generate SH, TH and higher optical harmonics with high efficiency.
HNPs markers are promising for applications in the field of bio -imaging due to the possibility of changing the wavelength of excitation, the high depth of imaging [10] and photostability for long-term observation. Such markers can be easily identified through their optical harmonics signals against the background due to the response of the components of biological tissues [11,12].
Typical HNPs are: KNbO3, BiFeO3 (BFO), LiNbO3, BaTiO3, Fe(IO3)3, Ba(BO2)2 (BBO), KTiOPO4 (KTP), and ZnO [13]. In general, any NPs based on already known crystals – frequency converters of laser radiation, can be used as HNPs. Compared to typical fluorophore molecules, HNPs have a much narrower spectral response (see Fig. 1.3) and the harmonic signals are large enough to be detected in most of the standard nonlinear optical microscopy systems (see Fig. 1.4).
Fig. 1.3 An example of the possibility of tuning the narrow peaks of the SHG signals: (1) HNPs Sr0.6Ba0.4Nb2O6 with a size of 42 nm in comparison with the luminescence bands of (2) CdSe quantum dots with the size of 4 nm; (3) upconversion NPs NaYF4:Er3+, Yb3+ with size of 18 nm under two-photon excitation [14].
Generally, an efficient generation of optical harmonics is obtained in nonlinear optical crystals under the conditions of phase synchronism or phase matching. Due to the strong dispersion of the refractive index, such conditions can be fulfilled in a NLO crystal only in a limited range of excitation wavelengths, for specific crystal orientations and polarizations of the pump beam. However, if the size of the crystals does not exceed the coherent length (typically below one micron), these restrictions are removed [15].
Optical contrast based on the frequency doubling and tripling process can bring a number of benefits to bioimaging. First of all, as only virtual electronic states are involved, there is limited energy absorption, preventing bleaching, which is usually observed in fluorescent probes. Thus, it is possible to achieve observations over a long period of time without reducing the signal intensity [11]. In addition, SHG and THG are non-resonant processes that can occur for any wavelength of excitation [10,11,19]. Consequently, the pump wavelength can be tuned to a specific spectral range, where the absorption and scattering of biological tissues are low, which limits photo-degradation and increases the depth of penetration [20]. The correct selection of pump wavelength also avoids overlays with auto-fluorescence of the sample and thus increases the contrast of the image. Moreover, the SH signal from NPs has a quadratic dependence on the intensity of the excitation. Detection and visualization of such individual biological markers typically require high peak excitation power, which is now easy to achieve with femtosecond laser sources, even at very moderate pulse energies [20,21].
In nonlinear microscopy, bleaching and blinking are well-known causes of image degradation. In the case of fluorescent probes, that reached the excited state when photons are absorbed, they can, with a negligible probability, pass on to another non-radiation-emitted excited state. Similarly, fluctuations in the intensity affecting radiation from quantum dots (blinking), as a consequence of capturing photo-excited electrons on the surface of HNPs, cannot occur with SHG and THG.
It is known that in the electrodipole approximation, the intensity of the second optical harmonic increases as the square of the nanocrystal volume. This was confirmed in the literature for various noncentrosymmetric oxide nanocrystals [22–24]. When the particles size decreases, there is thus a significant reduction of the generation efficiency (NPs with two times smaller radius should generate 64 times lower SHG signal) that imposes restrictions for practical applications. HNPs have many attractive characteristics, but real biological implementation sometimes requires a decrease of the sizes of the NPs for the best permeability in biological tissues and their successful natural removal from the body [25,26]. Particles with dimensions less than 10 nm [25] may be interesting for some specific applications. In that case, harmonic generation efficiency has to be increased.
One approach for that is to combine a dielectric oxide core with a metal shell to amplify local fields through resonance excitation of surface plasmons. Among such materials, it is worth mentioning BaTiO3 NPs with a gold shell, which demonstrated high SHG [27] or parametric amplification of light radiation [28]. A similar approach was observed for third harmonic generation of individual NPs located within a metallic nanoantenna gap that promotes the amplification of the local fields [29,30].
Generation of resonantly amplified SH is also well studied in pure metallic NPs [31], where the amplification of a local electric field is achieved thanks to the contribution of plasmon resonances. Such nanostructures generate a NLO response due to surface effects or disturbances of the symmetry of excited modes [32], since the bulk second-order susceptibility tensor χ(2) is zero due to the lattice’s symmetry.
In recent years, nanophotonics have been intensively developed on the basis of all-dielectrics nanostructures as an alternative to plasmonics. This approach is based on the use of bulk resonances of the Mie-type – resonances of displacement currents – instead of the response of surface plasmons. In accordance with the Mie theory of scattering, the resonance [33] is located at λ ~ nd, where d is the characteristic size of the NP and n its refractive index. Although the field amplification in these dielectric structures is usually weaker than that of metallic analogs, their high Q factor factors allow to implement effective NLO responses. Nonlinear effects, such as SHG and THG, are also sensitive to resonant properties of dielectric nanostructures. This allowed the researchers to observe a significant gain in SHG and THG efficiency in dielectric and semiconductor materials [34,35].
Fig. 1.4 An example of the simultaneous generation of signals of SH and TH by BiFeO3 harmonic NPs in stem cells of human skeletal muscle [36].
Another approach to extending the range of application of harmonic nanoparticles is to create multifunctional nanoprobes that combine generation of optical harmonics at pulsed excitation and up-conversion with continuous laser excitation [37]. Unlike traditional fluorescent markers with UV excitation (quantum dots, fluorophore molecules), upconverting nanoparticles allow pumping in the near-infrared range that can significantly reduce the contribution of tissue autofluorescence and increase imaging penetration in biological tissues up to 1 mm [36]. These new multifunctional nanoprobes combine resonant and nonresonant excitation conditions [38] with up-conversion and SHG processes and excitation in the first infrared window of biological tissues (700-900 nm), which extends the scope of applications of HNPs.
In the last few years, there has been a significant expansion of the use of HNPs, that requires a proper characterization of their nonlinear optical properties. At the beginning, the main attention was concentrated on the study of second harmonic generation, but over time, for more advanced applications, the response at the third and higher optical harmonics become relevant. Indeed, measuring the signals of several harmonics at the same time and comparing these images can significantly improve the accuracy of the identification of the HNPs against the background from biological tissues.
In reference [36], BFO HNPs was used as a marker for stem cells from human skeletal muscle (hMuStem) to study the feasibility of a therapeutic implementation of stem cell-based approaches, namely, their distribution and fixation. It was shown that the simultaneous measurement of SHG and THG allows to clearly identify the response from the HNPs against the response of biological tissues, for a significant increase in the selectivity of the images (see Fig. 1.4). The possibility of detecting 100 nm HNPs in muscle tissues, at a distance of more than 1 mm and with excitation in the second transparency window of the near-infrared range 1000 -1700 nm, has been demonstrated. These studies were performed for 14 days without any modification of the proliferative and morphological features of hMuStem cells. Other studies have also shown high biocompatibility of such NPs and efficiency for bioimaging applications [36].
Following these developments, it was then of particular interest to develop a quantitative method to measure the THG efficiency of the HNPs. From the experimental point of view, this measure is more complex compared to SHG.
One approach is to analyze harmonic signals from individual nanoparticles with multiphoton microscopy. However, it takes a lot of time for collecting a set of statistical data, and the results are difficult to analyze because NPs orientation is unknown [39–41].
For the study of SHG efficiency, as an alternative to microscopy, the hyper-Rayleigh scattering technique (HRS@SH) has been developed. HRS@SH is an experimental method based on the analysis of the scattered SH signal from a solution of molecules, which was then extended to the study of nanoparticles in colloidal suspensions [18-19]. Using this experimental technique, one can measure the orientation averaged second-order susceptibility of harmonic nanoparticles. The protocol is based on the comparison of the response from a colloidal suspension with the response from a reference solution of molecules [42–44], but applying this approach at the TH frequency(HRS@ TH) is significantly more complicated and less precise from the experimental point of view.
In this work, an experimental method based on the interface scanning technique at third harmonic (IS@TH)[45], that will be described in the experimental section, was optimized to study HNPs. We indeed applied this experimental method (see chapter 3) to derive the averaged third-order susceptibility of ZnO nanoparticles and compared the obtained values with Third Harmonic Scattering measurements (HRS@TH). Initially, the interface scanning method has been developed to derive the third harmonic susceptibilities of liquids and gases [46].
Moreover, a linear scattering and laser beam self-action effect were also applied to study ZnO harmonic nanoparticles or more complex nanostructures (see chapter 4). Finally, nonlinear optical properties of ZnO harmonic nanoparticles were also studied using a multiphoton microscopy setup (see chapter 3), that was developed during my last year of PhD program at the SYMME laboratory.
Composite materials based on KDP single crystals with incorporated inorganic NPs and organic impurities
Single crystals of potassium dihydrogen phosphate, known as KDP (KH2PO4), deuterated KDP (DKDP or KD*P), and ammonium dihydrogen phosphate, known as ADP (NH4H2PO4) groups are widely used in nonlinear optics, optoelectronics and laser engineering. The high laser damage threshold and the possibility of growing crystals of large size makes these materials economically beneficial for the production of wide-aperture laser frequency converters and Pockels cells for powerful laser setups [47]. In KDP crystals, there is a large amount of hydrogen bonds, so they are able to effectively capture
impurities of different nature [48], making them model objects for the search for new memory elements, substances for nonlinear optics and laser [49].
Fig. 1.5 Scheme of intrinsic defects energy levels (a) of nanocrystalline TiO2 in the anatase modification [50,51] and (b) KDP single crystal [52]. For the anatase, the scheme of excitation/relaxation of carriers under the influence of laser radiation with quantum energies 1.17 eV (1064 nm) and 2.33 eV (532 nm) is given. VB and CB – valence band and conduction bands, ST and DT – shallow and deep traps. (c) Photo of KDP single crystals: TiO2 with incorporated anatase NPs, TEM image of the NPs, and the structure of their layered entry into the KDP matrix.
However, due to the relatively low nonlinear coefficients of KDP, on practice it is more efficient to use different composites based on KDP matrix as “host” with different “guest” inclusions such as organic molecules or nanoparticles. The influence of “guest” subsystems on “host” matrix will be described in literature review.
It is known that the doping of KDP crystals with complex organic molecules such as L-arginine leads to an increase of the efficiency of the second harmonic generation by 30-70%[53]. Doping KDP crystals with urea (carbamide) at a concentration of 2 wt. % leads to an increase of the efficiency of the SHG by 30% [2]. This is due to the additional deformation of the charged tetrahedron (PO4)3- due to hydrogen bonds formation between the amino group of urea and the hydrophosphate (H2PO4) group of the crystalline matrix.
It was also shown that the incorporation of metal oxides nanoparticles into a crystalline matrix of nominally pure KDP leads, under certain conditions of excitation, to a amplification of the cubic NLO response [54], that is induced by a nonlocal transformation of the hydrogen bonds system around the NPs in the crystal [55]. The dimensions of the formed nanosized domains induced by NPs spontaneous polarization substantially increase the size of the NPs itself. As a result, at room temperature, the formation of an effective medium of « guest-host » type occurs, where the crystalline KDP matrix in the paraelectric state acts as the « host », and in the role of the « guest » – the NPs surrounded by a nonlocal layer of permeated hydrogen bounds of the KDP matrix. This leads to an increase in photoinduced changes of the macroscopic refractive index, leading to self-focusing phenomenon, which finally causes an increase of optical harmonics generation efficiency.
Table of contents :
Introduction
Chapter 1 Literature review
1.1 Harmonic nanoparticles
1.2 Composite materials based on KDP single crystals with incorporated inorganic NPs and organic impurities
Chapter 2 Experimental optical techniques
2.1 Elastic light scattering indicatrices technique
2.2 Nonlinear optical response of medium
2.3 Characterization of the nonlinear susceptibilities of HNPs Conclusions to the chapter 2
Chapter 3 Nonlinear optical response of ZnO harmonic nanoparticles
3.1 Preparation of ZnO colloidal suspensions
3.2 Averaged third-order susceptibility from the interface scanning technique
3.3. Multiphoton microscopy of single ZnO nanoparticles Conclusions to the chapter 3
Chapter 4 Characterization of different types of ZnO based nanostructures and bulk crystals
4.1 Studied materials
4.2 Influence of the synthesized ZnO NPs size on the nonlinear optical response
4.3 Multiphoton microscopy of raspberry-like ZnO NPs
4.4 Analysis of the energy structure and nonlinear optical response of bulk ZnO crystals with different content of intrinsic defects
Conclusions to the chapter 4
Chapter 5 Analysis of the optical harmonics generation efficiency of composite materials based on single crystals of KDP
5.1 Composites based on KDP single crystals
5.2 Nonlinear optical response of KDP single crystal with incorporated TiO2 nanoparticles
5.3 Influence of Al2O3·nH2O nanofibers incorporation in KDP single crystal on the nonlinear optical response
5.4 Influence of KDP single crystals doping by L-arginine on NLO response efficiency
Conclusions to the chapter 5
General Conclusion
Publications list
Appendix
References
Résumé en français
