This PDF file contains the front matter associated with SPIE Proceedings Volume 12394, including the Title Page, Copyright information, Table of Contents, and Committee information.

In this paper a wavelength-multiplexing based super-resolving concept is presented to allow high resolution imaging through blood. We use temporally pulsed and spectrally wide-band laser while at its output we have special grating and a spatial 2-D transmission mask allowing to project wavelengths’ dependent high-resolution spatial orthogonal encoding patterns (different spatial patterns for each wavelength). The ballistic photons of the short temporal pulses allow the high-resolution encoding pattern to reach the inspected object through the scattering blood medium without being spatially blurred. The light is intentionally collected via low resolution optics. The high-resolution reconstruction can be obtained digitally by post processing or optically by passing the collected low-resolution data through a similar grating and 2-D mask which do an all-optical decoding. After summing all the images together, the super-resolved reconstruction through the highly scattering blood medium is formed.

Various plasmonic nanostructure-based substrates are used to detect biological signals beyond the diffraction limit with a high signal-to-noise ratio. These approaches take advantage of excitation of localized surface plasmon to acquire high-frequency biological signals while preserving photon energy. Numerous techniques, including focused ion beam, electronbeam lithography, and reactive ion etching, have been used to fabricate plasmonic substrates. However, these fabrication techniques are time and resource-consuming. In contrast, disordered nanostructure-based substrates have attracted interests due to the easy fabrication steps and potential cost savings. Metallic nanoisland substrates, for instance, can be mass-produced using thin film deposition and annealing without lithographic process. In this work, we have investigated nanospeckle illumination microscopy (NanoSIM) using disordered near-field speckle illumination generated by nanoisland substrate. Selectively activated fluorescence wide-field images were obtained by nanospeckle illumination generated on the nanoisland substrate. Super-resolved fluorescence images were reconstructed by an optimization algorithm based on blind structured illumination microscopy. Experimental studies of various biological targets including HeLa cell membranes were performed to demonstrate the performance of NanoSIM. Using NanoSIM, we were able to improve spatial resolution of ganglioside distribution in HeLa cells targeted by CT-B by more than threefold compared to the diffraction-limited images. Note that the accessibility of super-resolution imaging techniques can be enhanced by the nanospeckle illumination of disordered metallic nanoislands. These results may be used in imaging and sensing systems that work with detecting biological signals beyond diffraction limits in various applications.

Imaging in opaque media is limited in spatial resolution. Therefore, detection of nanoelements that are behind the imaging resolution limit is challenging. In this work, we used the iterative multiplane optical properties extraction (IMOPE) technique to detect nanoparticles (NP) within the different skin layers. The IMOPE technique was originally designed for detecting NPs in turbid environment based on their scattering property. However, the scattering depends on the size of the particle, therefore detecting extremely small particles, with negligible scattering, was still a challenge for the IMOPE technique. In our work, we adapted the IMOPE technique to detect nanodiamonds (ND) as such, with negligible scattering but nonnegligible absorption, inside the different layers of the skin. In the following manuscript we present an extension of the automated technique, that is more robust and applicable to non-symmetrical samples.

Imaging with conventional optical systems has the limitation of the diffraction limit of the point spread function (PSF) where the maximum attainable resolution reaches down to half of the incident wavelength in lateral direction and down to the wavelength in axial direction. In fact, the tradeoff between the lateral and axial confinement is inevitable resulting in a deterioration of the 3D optical image. In this work, we report a metalens design attaining high resolution imaging in lateral and axial directions at 𝜆=3𝜇m. The proposed design employs a 2D square photonic crystal lattice of silicon nanorods with an engineered optical cavity and a silver slit on top of the structure. Our design exhibits a simultaneous sub-diffraction limit in lateral and axial directions reaching down to 0.38λ and to 0.8λ, respectively. The ideal PSF requires the equality of the full width at half maximum (FWHM) for both lateral and axial directions. Hence, the figure of merit (FOM) can be expressed as the ratio between the (FWHM) in lateral and axial directions approaching unity for ideal PSF. The proposed design demonstrates enhanced FOM, up to 0.47, compared to the confocal microscopy that shows FOM of 0.36. Our proposed design provides a great potential usage in the field of biomedical imaging and molecular dynamics.

In 2013 we proposed the magneto-thermo-acoustic effect [Piao et al, Med. Phys. 2013], which refers to acoustic emission from magnetic nanoparticles (MNPs) when thermally mediated under an alternating magnetic field (AMF) applied in a pulsed or frequency-chirped mode. Several independent experimental studies have since validated magneto-thermo-acoustic effect in association with various modes of AMF application including pulsing and chirping of the intensity modulation [Feng et al, Appl Phys. Lett., 2013]. Later Kellnberger et al demonstrated that [Phys Rev Lett. 2016] a continuous-wave (CW) AMF applying to MNPs activated acoustic emission at ONLY the second-harmonic frequency of AMF. We predict that applying a static bias magnetic field to MNPs in the presence of a CW AMF produces acoustic emission at the fundamental frequency of AMF, in addition to the known second harmonic frequency of AMF. The mechanism of this dual-frequency acoustic emission is projected as the partial magnetization of MNPS by the bias field affecting the AMF mediation of MNPs. The coupling between the two magnetic fields in modulating the magnetic susceptibility of MNPs causes acoustic emission at both the fundamental and second harmonic frequencies of AMF. Interestingly, the intensity ratio of the acoustic emission at the two frequencies is determined uniquely by the intensity ratio of the two magnetic fields. This spectral intensity characteristic of the dual-frequency acoustic emission from MNPs can thus be made spatially unique by controlling the bias field over AMF. This potentiates spatial encoding or magnetically scanned imaging of MNPs towards theragnostic applications.

Environmental pollution by microplastics (MPs) represents a serious burden of the 21st century. Sensing the interactions of photosynthetic organisms with MPs is based on the study of their endogenous fluorescence derived from chlorophylls. Fluorescently labelled custom-made MPs were tested. We also recorded endogenous fluorescence of the moss in the presence of “naturally-occurring” MPs (polyethylene content of 2 mg/g, non fluorescent) in suspended matter (SM) from the river Rhine. Performed experiments evaluated the distribution of the MPs, as well as the sensitivity of endogenous fluorescence of chlorophylls to their presence. Understanding the interaction of living organisms with MPs will help to assess the impact of this environmental pollution and eventually to propose new approaches for its removal from water sources.

The unique fluorescent nanomaterials known as carbon dots (CDs) are highly resistant to photobleaching, have low toxicity, and are well soluble in water. Polyethyleneimine (PEI) coated CDs are a novel fluorophore with good biocompatibility and pH sensing ability. Here, p-phenylenediamine (p-PD) is used as a carbon source and hyperbranched PEI is used as a surface passivation agent in a simple, one-step hydrothermal synthesis process. The CDs optical characteristics are pH-responsive due to the presence of different amine groups on PEI, which is functional polycationic polymer. The limits of techniques based on fluorescence intensity can be overcome by fluorescent lifetime imaging microscopy (FLIM), a very sensitive method for detecting a microenvironment. In this study, FLIM was used to measure pH with pH-sensitive CDs. These molecules are nontoxic to the cells, and the positively charged CDs have the potential for nuclear targeting, allowing for electrostatic contact with DNA in the nucleus. Higher wavelengths have a larger penetration depth of electromagnetic radiation and low tissue autofluorescence, hence CDs emitting at these wavelengths are used for biolabeling applications. However, the quantum yield of these synthesized red-emissive CDs is lower. In order to enhance it, they are conjugated with gold nanoparticles(AuNPs) for metal enhanced fluorescence (MEF). Through a potent covalent bond between them, the AuNPs are linked to CDs surfaces. These gold-CDs nanoconjugate can be used in the future for targeted imaging applications.

Although single point time-resolved fluorescence anisotropy (FA) measurements are well established and routinely used for various applications in many laboratories, only a few reports described their extension into two-dimensional (2D) time-resolved FA imaging (TR-FAIM). The ability to perform TR-FAIM can offer cellular imaging based on the rotational correlation time (θ) that depends on the viscosity and dynamic properties of the tissues. We extended existing frequency domain (FD) fluorescence lifetime (FLT) imaging microscopy (FLIM) to FD TR-FAIM, which produces visual maps of θ. The proof of concept of the FD TR-FAIM was validated on 7 fluorescein solutions with increasing viscosities (achieved by increasing glycerol concentration between 0-80%). The studies were performed using images of θ as well as by characterizing the peak (mode) and the full width half maximum (FWHM) of its histograms (of normal probability distribution) and extracting the limiting FA (r₀). The θ of the 7 solutions was significantly increased from 0.15±0.05 to 11.25±1.87ns, whereas r₀ decreased from 0.40±0.01 to 0.30±0.06. The FD TR-FAIM provides wide-field imaging of the θ of the fluorophore, and hence offers a potential simultaneous interrogation with great sensitivity of diverse chemical and physical phenomena. In addition, as θ can vary according to the local microenvironment and across the sample under investigation, it can characterize different compartments of complex structures such as cells. Through the FD TR-FAIM a large variety of information can be probed from each sample and therefore it may become a reliable and powerful diagnostic tool for cellular imaging and biosensing.