Biological Transparency Window Research Articles

Heat, pH, and salt: synthesis strategies to favor formation of near-infrared emissive DNA-stabilized silver nanoclusters.

We present chemical synthesis strategies for DNA-stabilized silver nanoclusters (AgN-DNAs) with near-infrared (NIR) emission in the biological tissue transparency windows. Elevated temperatures can significantly increase chemical yield of near-infrared nanoclusters. In most cases, basic pH favors near-infrared nanoclusters while micromolar amounts of NaCl inhibit their formation.

Electron count and ligand composition influence the optical and chiroptical signatures of far-red and NIR-emissive DNA-stabilized silver nanoclusters.

Near-infrared (NIR) emissive DNA-stabilized silver nanoclusters (AgN-DNAs) are promising fluorophores in the biological tissue transparency windows. Hundreds of NIR-emissive AgN-DNAs have recently been discovered, but their structure-property relationships remain poorly understood. Here, we investigate 19 different far-red and NIR emissive AgN-DNA species stabilized by 10-base DNA templates, including well-studied emitters whose compositions and chiroptical properties have never been reported before. The molecular formula of each purified species is determined by high-resolution mass spectrometry and correlated to its optical absorbance, emission, and circular dichroism (CD) spectra. We find that there are four distinct compositions for AgN-DNAs emissive at the far red/NIR spectral border. These emitters are either 8-electron clusters stabilized by two DNA oligomer copies or 6-electron clusters with one of three different ligand compositions: two oligomer copies, three oligomer copies, or two oligomer copies with additional chlorido ligands. Distinct optical and chiroptical signatures of 6-electron AgN-DNAs correlate with each ligand composition. AgN-DNAs with three oligomer ligands exhibit shorter Stokes shifts than AgN-DNAs with two oligomers, and AgN-DNAs with chlorido ligands have increased Stokes shifts and significantly suppressed visible CD transitions. Nanocluster electron count also significantly influences electronic structure and optical properties, with 6-electron and 8-electron AgN-DNAs exhibiting distinct absorbance and CD spectral features. This study shows that the optical and chiroptical properties of NIR-emissive AgN-DNAs are highly sensitive to nanocluster composition and illustrates the diversity of structure-property relationships for NIR-emissive AgN-DNAs, which could be harnessed to precisely tune these emitters for bioimaging applications.

Monitoring the Activity and Inhibition of Cholinesterase Enzymes using Single-Walled Carbon Nanotube Fluorescent Sensors.

Cholinesterase enzymes are involved in a wide range of bodily functions, and their disruption is linked to pathologies such as neurodegenerative diseases and cancer. While cholinesterase inhibitors are used as drug treatments for diseases such as Alzheimer and dementia at therapeutic doses, acute exposure to high doses, found in pesticides and nerve agents, can be lethal. Therefore, measuring cholinesterase activity is important for numerous applications ranging from the search for novel treatments for neurodegenerative disorders to the on-site detection of potential health hazards. Here, we present the development of a near-infrared (near-IR) fluorescent single-walled carbon nanotube (SWCNT) optical sensor for cholinesterase activity and demonstrate the detection of both acetylcholinesterase and butyrylcholinesterase, as well as their inhibition. We show sub U L–1 sensitivity, demonstrate the optical response at the level of individual nanosensors, and showcase an optical signal output in the 900–1400 nm range, which overlaps with the biological transparency window. To the best of our knowledge, this is the longest wavelength cholinesterase activity sensor reported to date. Our near-IR fluorescence-based approach opens new avenues for spatiotemporal-resolved detection of cholinesterase activity, with numerous applications such as advancing the research of the cholinergic system, detecting on-site potential health hazards, and measuring biomarkers in real-time.

Super‐Resolution Near‐Infrared Fluorescence Microscopy of Single‐Walled Carbon Nanotubes Using Deep Learning

Single‐walled carbon nanotubes (SWCNTs) have unique optical and physical properties, with numerous biomedical imaging and sensing applications, owing to their near‐infrared (nIR) fluorescence which overlaps with the biological transparency window. However, their longer emission wavelengths compared to emitters in the visible range result in a lower resolution due to the diffraction limit. Moreover, the elongated high‐aspect‐ratio structure of SWCNTs poses an additional challenge on super‐resolution techniques that assume point emitters. Utilizing the advantages of deep learning and convolutional neural networks, along with the super‐resolution radial fluctuation (SRRF) algorithm for network training, a fast, parameter‐free, computational method is offered for enhancing the spatial resolution of nIR fluorescence images of SWCNTs. An average improvement of 22% in the resolution and 47% in signal‐to‐noise ratio (SNR) compared to the original images is shown, whereas SRRF leads to only 24% SNR improvement. The approach is demonstrated for a variety of SWCNT densities and length distributions, and a wide range of imaging conditions with challenging SNRs, including real‐time videos, without compromising the temporal resolution. The results open the path for accelerated and accessible super‐resolution of nIR fluorescent SWCNTs images, further advancing their applicability as nanoscale optical probes.

Plasmonically enhanced two-photon absorption induced photoacoustic microscopy with laser-synthesized TiN nanoparticles

Combining photonic excitation and acoustic detection, photoacoustic imaging (PAI) presents one of the most promising noninvasive biomedical diagnostic modalities, but this technique still lacks efficient nano-sized contrast agents absorbing light in the region of relative tissue transparency (630–900 nm). Here, we explore the use of titanium nitride (TiN) nanoparticles (NPs) fabricated by methods of pulsed laser ablation in liquids as a contrast agent in PAI. When prepared in acetone, the NPs are spherical, have an average size of 25 nm, and exhibit a broad plasmonic absorption peak around 700 nm. We show that solutions of these NPs render possible a strong nonlinear photoacoustic response and the generation of photoacoustic images with 67 μm resolution within the biological transparency window. The observed effect is explained by a plasmonically enhanced two-photon absorption process in TiN NPs. Combined with earlier demonstrated capability of generating photothermal therapeutic effect, relative chemical purity, and excellent biocompatibility, laser-synthesized TiN NPs promise attractive applications in biomedical theranostics involving imaging modalities based on photoacoustics microscopy or tomography.

Magnetic Field-Modulated Plasmonic Scattering of Hybrid Nanorods for FFT-Weighted OCT Imaging in NIR-II.

We report a method for fast Fourier transform (FFT)-weighted optical coherence tomography (OCT) in the second biological tissue transparency window by actively modulating the plasmonic scattering of Fe3O4@Au hybrid nanorods using magnetic fields. Instead of tracking the nanoparticles' lateral displacement in conventional magnetomotive OCT imaging, we monitor the nanorod rotation and optical signal changes under an alternating magnetic field in real time. The coherent rotation of the nanorods with the field produces periodic OCT signals, and the FFT is then used to convert the periodic OCT signals in the time domain to a single peak in the frequency domain. This allows automatic screening of nanorod signals from the random biological noises and reconstruction of FFT-weighted images using a computer program based on a time-sequence image set. Compared with conventional magnetomotive OCT, the FFT-weighted imaging technique creates enhanced OCT images with dB-scale contrast over an order of magnitude higher than the original images.

Germanium Nanoparticles Prepared by Laser Ablation in Low Pressure Helium and Nitrogen Atmosphere for Biophotonic Applications.

Due to particular physico-chemical characteristics and prominent optical properties, nanostructured germanium (Ge) appears as a promising material for biomedical applications, but its use in biological systems has been limited so far due to the difficulty of preparation of Ge nanostructures in a pure, uncontaminated state. Here, we explored the fabrication of Ge nanoparticles (NPs) using methods of pulsed laser ablation in ambient gas (He or He-N2 mixtures) maintained at low residual pressures (1–5 Torr). We show that the ablated material can be deposited on a substrate (silicon wafer in our case) to form a nanostructured thin film, which can then be ground in ethanol by ultrasound to form a stable suspension of Ge NPs. It was found that these formed NPs have a wide size dispersion, with sizes between a few nm and hundreds of nm, while a subsequent centrifugation step renders possible the selection of one or another NP size fraction. Structural characterization of NPs showed that they are composed of aggregations of Ge crystals, covered by an oxide shell. Solutions of the prepared NPs exhibited largely dominating photoluminescence (PL) around 450 nm, attributed to defects in the germanium oxide shell, while a separated fraction of relatively small (5–10 nm) NPs exhibited a red-shifted PL band around 725 nm under 633 nm excitation, which could be attributed to quantum confinement effects. It was also found that the formed NPs exhibit high absorption in the visible and near-IR spectral ranges and can be strongly heated under photoexcitation in the region of relative tissue transparency, which opens access to phototherapy functionality. Combining imaging and therapy functionalities in the biological transparency window, laser-synthesized Ge NPs present a novel promising object for cancer theranostics.

Core-multishell lanthanide-doped nanocomposite by one-pot synthesis for NIR-II emissions-based temperature sensing

The performances of nanothermometers based on visible light are inevitably degraded in biological applications because visible light is greatly weakened after passing through tissues. Herein, in view of the second biological optical transparency window, a type of tailored core-multishell lanthanide-doped nanocomposite was synthesized with facile one-pot coprecipitation method to supplant complex method demanding stepwise thermal decomposition. Proved to have excellent dispersibility, crystallinity, stability and luminous efficiency, the nanocomposite is capable of enhanced NIR-II emissions and a broad temperature detection scope based on three kinds of spectral parameters relevant to its unique core-multishell structure. The maximum absolute and relative sensitivity of luminescence intensity ratios in low temperature range can reach up to 11.3% K−1 and 1.27% K−1, respectively, and a theoretical derivation is provided to address the peculiar V-shaped trend of NIR-II luminescence lifetime. The prepared nanocomposite can serve as a promising candidate for optical thermometer in the fields of cryoablation of diseased tissues and industrial applications in strong magnetic or radiofrequency fields.

Two-Photon Absorption: An Open Door to the NIR-II Biological Window?

The way in which photons travel through biological tissues and subsequently become scattered or absorbed is a key limitation for traditional optical medical imaging techniques using visible light. In contrast, near-infrared wavelengths, in particular those above 1000 nm, penetrate deeper in tissues and undergo less scattering and cause less photo-damage, which describes the so-called “second biological transparency window”. Unfortunately, current dyes and imaging probes have severely limited absorption profiles at such long wavelengths, and molecular engineering of novel NIR-II dyes can be a tedious and unpredictable process, which limits access to this optical window and impedes further developments. Two-photon (2P) absorption not only provides convenient access to this window by doubling the absorption wavelength of dyes, but also increases the possible resolution. This review aims to provide an update on the available 2P instrumentation and 2P luminescent materials available for optical imaging in the NIR-II window.

Modulation Technique of Localized Surface Plasmon Resonance of Palladium Nanospheres by Coating with Titanium Dioxide Shell for Application to Photothermal Therapy Agent

Although plasmonic palladium (Pd) nanospheres are thermodynamically stable and have high photothermal conversion due to the free and bound electron coupling associated with the intrinsic high interband transition, they have not attracted attention as a photothermal conversion material for next-generation photothermal cancer therapy. This is because the Pd nanospheres generate the localized surface plasmon resonance (LSPR) intrinsically in the ultraviolet region, which is far away from the biological transparent window (750–900 nm). In this study, we controlled the LSP wavelength of Pd nanospheres by coating with high refractive index TiO2 shells taking advantage of the Pd LSPR which is highly sensitive to changes in the local refractive index around the nanospheres. Our calculations indicated that the absorption cross section at 808 nm (corresponding to the wavelength used for photothermal treatment) was increased by 4.5 times by redshifting the LSPR and increasing the extinction intensity associated with the coating with TiO2 shell. Experiments confirmed the theoretical prediction in that the LSPR of the synthesized Pd nanospheres with a diameter of 81 nm was significantly redshifted by coating with amorphous TiO2 shell, resulting in significant large extinction intensity at 808 nm. The photothermal conversion efficiency was estimated to be 50%. In vitro cell tests, HeLa cells incubated with 100–300 μg/mL TiO2-coated Pd nanospheres were efficiently killed by irradiating 808 nm laser (1.8 W) even though the nanospheres with the same concentrations showed little cytotoxicity. These results indicate that the Pd nanospheres coated with high refractive index shells can be promising as a photothermal therapy agent.

Highly efficient red-emitting carbon dots as a “turn-on” temperature probe in living cells

Nanothermometers, which can precisely detect the intracellular temperature changes, have great potential to solve questions concerning the cellular processes. Thus, the temperature sensors that provide fluorescent “turn-on” signals in the biological transparency window are of highly desirable. To meet these criteria, this work reported a new “turn-on” carbon dot (CD)-based fluorescent nanothermometry device for sensing temperature in living cells. The CDs that emit bright red fluorescence (R-CDs; λmax = 610 nm in water) were synthesized with o-phenylenediamine as carbon precursor via a facile solvothermal method. The R-CDs in water were almost nonfluorescent at 15 °C. As the temperature increased, the fluorescence intensity of R-CDs exhibited a gradual increase and the final enhancement factor was greater than 21-fold. The fluorescence intensity exhibited a linear response to temperature and a high-sensitive variation of ≈13.3 % °C−1 was detected within a broad temperature range of 28–60 °C. Moreover, the R-CD thermal sensors also exhibited high storage stability, excellent response reversibility and superior photo- and thermo-stability. Due to its good biocompatibility and “intelligent” response to external temperature, the nanothermometer could be applied for sensing temperature changes in biological media.

Uni- and multivariate calibration of tempe­rature from the neodymium fluorescence spectra in nanocrystals of yttrium-gadolinium oxide and yttrium-gadolinium garnet

The use of neodymium-doped nanocrystalline powders of yttrium-gadolinium oxide and yttrium gadolinium garnet to increase the sensitivity of local fluorescent optical temperature sensors is considered. Based on the temperature dependences of the neodymium fluorescence spectra in this powders, univariate (using fluorescence intensity ratio from thermally coupled energy levels of the activator) and multivariate (using the partial least squares method) calibration models are developed. When using the spectral range 860 – 950 nm falling into the first biological transparency window (700 – 980 nm), both calibration models have a standard deviation of about 10 % and are comparable in accuracy. The spectral variables selection by searching combination moving window in the multivariate model made it possible to reduce the root mean square error for yttrium-gadolinium oxide by more than 12 times (from 9.8 to 0.8 °C), and for yttrium-gadolinium garnet by more than 2 times(from 8.7 to 4.0 °С). The result obtained indicatesthe proposed neodymium-doped nanocrystalline powders and multivariate methods of calibration can be used to localise areas with febrile temperatures for biological and medical purposes.

Plasmon-enhanced near-infrared fluorescence detection of traumatic brain injury biomarker glial fibrillary acidic protein in blood plasma

An ultrasensitive plasmonic near-infrared fluorescent biosensor substrate has been developed for detection of glial fibrillary acidic protein (GFAP) biomarker in blood plasma, an important protein biomarker of traumatic brain injury (TBI). To minimize the interference from blood plasma sample matrix, a near-infrared fluorophore in the first biological transparency window is used in the biosensor. To amplify the fluorescence signals, a plasmonic gold nanopyramid array has been coupled to the fluorophore. Finite-difference time-domain simulation reveals that the excitation enhancement is primarily responsible for the fluorescence enhancement owing to the intense local electric field excited on the corners and edges. As a result, this biosensor exhibits a lower limit of detection of 0.6 pg/mL toward detection of GFAP in blood plasma.

Super-Resolution Radial Fluctuations (SRRF) nanoscopy in the near infrared.

Super resolution microscopy methods have been designed to overcome the physical barrier of the diffraction limit and push the resolution to nanometric scales. A recently developed super resolution technique, super-resolution radial fluctuations (SRRF) [Nature communications, 7, 12471 (2016)10.1038/ncomms12471], has been shown to super resolve images taken with standard microscope setups without fluorophore localization. Herein, we implement SRRF on emitters in the near-infrared (nIR) range, single walled carbon nanotubes (SWCNTs), whose fluorescence emission overlaps with the biological transparency window. Our results open the path for super-resolving SWCNTs for biomedical imaging and sensing applications.

Visible-light and near-infrared fluorescence and surface-enhanced Raman scattering point-of-care sensing and bio-imaging: a review.

This review article deals with the concepts, principles and applications of visible-light and near-infrared (NIR) fluorescence and surface-enhanced Raman scattering (SERS) in in vitro point-of-care testing (POCT) and in vivo bio-imaging. It has discussed how to utilize the biological transparency windows to improve the penetration depth and signal-to-noise ratio, and how to use surface plasmon resonance (SPR) to amplify fluorescence and SERS signals. This article has highlighted some plasmonic fluorescence and SERS probes. It has also reviewed the design strategies of fluorescent and SERS sensors in the detection of metal ions, small molecules, proteins and nucleic acids. Particularly, it has provided perspectives on the integration of fluorescent and SERS sensors into microfluidic chips as lab-on-chips to realize point-of-care testing. It has also discussed the design of active microfluidic devices and non-paper- or paper-based lateral flow assays for in vitro diagnostics. In addition, this article has discussed the strategies to design in vivo NIR fluorescence and SERS bio-imaging platforms for monitoring physiological processes and disease progression in live cells and tissues. Moreover, it has highlighted the applications of POCT and bio-imaging in testing toxins, heavy metals, illicit drugs, cancers, traumatic brain injuries, and infectious diseases such as COVID-19, influenza, HIV and sepsis.

Efficient Triplet-Triplet Annihilation Upconversion in Solution and Hydrogel Enabled by an S-T Absorption Os(II) Complex Dyad with an Elongated Triplet Lifetime.

A new Os(II) complex dyad featuring direct singlet-to-triplet (S-T) absorption and intramolecular triplet energy transfer (ITET) with lifetime up to 7.0 μs was designed to enhance triplet energy transfer efficiency during triplet-triplet annihilation upconversion (TTA-UC). By pairing with 9,10-bis(phenylethynyl)anthracene (BPEA) as a triplet acceptor, intense upconverted green emission in deaerated solution was observed with unprecedented TTA-UC emission efficiency up to 26.3% (with a theoretical maximum efficiency of 100%) under photoexcitation in the first biological transparency window (650-900 nm). Meanwhile, a 7.1% TTA-UC emission efficiency was acquired in an air-saturated hydrogel containing the photosensitizer and a newly designed hydrophilic BPEA derivative. This ITET mechanism would inspire further development of a highly efficient TTA-UC system for biological fields and renewable energy production.

Lifetime of the 3H4 Electronic State in Tm3+-Doped Upconverting Nanoparticles for NIR Nanothermometry.

Emission bands from thermally coupled states in lanthanide-doped nanoparticles have been studied for ratiometric nanothermometry in biological applications. Unfortunately certain factors such as water absorption distort the intensity, limiting the accuracy of ratiometric nanothermometry. However, the decay time of such states does not suffer from such distortions. We introduce the decay time of the 3H4 state in Yb3+, Tm3+-doped nanoparticles for improved nanothermometry. The strong 800 nm upconversion emission exists in the first biological transparency window. This is the first use of a single upconversion band for lifetime nanothermometry.

Trimodal Ratiometric Luminescent Thermometer Covering Three Near-Infrared Transparency Windows.

Near-infrared (NIR) transparency windows have evoked considerable interest in biomedical thermal imaging owing to the superior tissue penetration and the high signal-to-noise ratio, allowing in vivo real-time temperature reading with nanometric spatial resolution. Here, we develop a multimode nonintrusive luminescent thermometer based on the Y3Al5O12 (YAG):Cr3+/Ln3+ (Ln = Ho, Er, Yb) phosphor, which covers three NIR biological transparency windows, enabling cross-checking readings with high sensitivity and a high penetration depth. Utilizing the energy transfer between lanthanide ions and transition-metal ions, the Cr3+/Ln3+-activated upconversion emissions provide ideal signals for ratiometric luminescent thermometry of the NIR-I mode. The phonon-assisted downshifting emissions of Er3+/Ho3+ are used to construct the NIR-III/II mode, and the NIR-III mode is based on the thermal coupling between stark levels of 4I13/2 (Er3+). Three independent modes show distinct thermometric performance in different NIR transparency windows and temperature ranges, and the combination of the three modes is conducive to obtain more accurate temperature readings in a broad temperature range, which paves the way toward versatile luminescent thermometers.

High-efficiency broadband achromatic metalens for near-IR biological imaging window

Over the past years, broadband achromatic metalenses have been intensively studied due to their great potential for applications in consumer and industry products. Even though significant progress has been made, the efficiency of technologically relevant silicon metalenses is limited by the intrinsic material loss above the bandgap. In turn, the recently proposed achromatic metalens utilizing transparent, high-index materials such as titanium dioxide has been restricted by the small thickness and showed relatively low focusing efficiency at longer wavelengths. Consequently, metalens-based optical imaging in the biological transparency window has so far been severely limited. Herein, we experimentally demonstrate a polarization-insensitive, broadband titanium dioxide achromatic metalens for applications in the near-infrared biological imaging. A large-scale fabrication technology has been developed to produce titanium dioxide nanopillars with record-high aspect ratios featuring pillar heights of 1.5 µm and ~90° vertical sidewalls. The demonstrated metalens exhibits dramatically increased group delay range, and the spectral range of achromatism is substantially extended to the wavelength range of 650–1000 nm with an average efficiency of 77.1%–88.5% and a numerical aperture of 0.24–0.1. This research paves a solid step towards practical applications of flat photonics.

Nd3+-Doped Lanthanum Oxychloride Nanocrystals as Nanothermometers.

The development of optical nanothermometers operating in the near-infrared (NIR) is of high relevance toward temperature measurements in biological systems. We propose herein the use of Nd3+-doped lanthanum oxychloride nanocrystals as an efficient system with intense photoluminescence under NIR irradiation in the first biological transparency window and emission in the second biological window with excellent emission stability over time under 808 nm excitation, regardless of Nd3+ concentration, which can be considered as a particular strength of our system. Additionally, surface passivation through overgrowth of an inert LaOCl shell around optically active LaOCl/Nd3+ cores was found to further enhance the photoluminescence intensity and also the lifetime of the 1066 nm, 4F3/2 to 4I11/2 transition, without affecting its (ratiometric) sensitivity toward temperature changes. As required for biological applications, we show that the obtained (initially hydrophobic) nanocrystals can be readily transferred into aqueous solvents with high, long-term stability, through either ligand exchange or encapsulation with an amphiphilic polymer.