This PDF file contains the front matter associated with SPIE Proceedings Volume 9339, including the Title Page, Copyright information, Table of Contents, Introduction (if any), and Conference Committee listing.

Understanding the structure and function of living cells from the nano to the micron scale remains a grand challenge. While X-ray diffraction and electron microscopy reveal nanoscale information from cellular structures, they operate with lifeless specimens. By contrast, optical techniques are well suitable for studying live cells. However, the resolution of the far-field “linear” microscopy is approximately 300 nm, a manifestation of Heisenberg’s uncertainty relation. Thus, pushing the biophotonics investigation toward the molecular scale is faced with significant challenges but also with unique opportunities. We will describe various principles dedicated to this goal and present some recent advances in phase sensitive measurements.

The use of the NIR spectral region (650-900 nm) for bioanalytical and biomedical analyses is advantageous due to the inherently lower background interference in biological matrices and the high molar absorptivities of NIR chromophores. There are several different groups of NIR fluorescing dye are available for bioanalytical applications. One of these groups, NIR carbocyanines are increasingly used in analytical, bioanalytical and medical applications. These dyes can be used as reporter labels for sensitive bioanalytical use, such as immunochemistry. Due to the spectroscopic sensitivity of NIR carbocyanines for polarity changes in the microenvironment fluorescence quantum yield can vary significantly dependent on the microenvironment. NIR dyes can have relatively low fluorescent quantum yields as compared to visible fluorophores, especially in aqueous buffers but the lower quantum yield is compensated for by a much higher molar absorptivity. The fluorescence intensity of NIR reporting labels can significantly be increased by enclosing several dye molecules in silica nanoparticles. Incorporation of NIR dyes in silica nanoparticles creates a unique challenge as these dyes can be unstable under certain chemical conditions present during silica nanoparticles syntheses. In addition, self quenching may also become a problem for carbocyanines at higher a concentrations that typically found inside of NIR dye loaded silica nanoparticles. Dyes possessing high Stokes’ shift can significantly reduce this problem. NIR carbocyanines are uniquely positioned for achieving this goal using a synthetic route that substitutes meso position halogens in NIR fluorescent carbocyanines with a linker containing amino moiety, which can also serve as a linker for covalently attaching the dye molecule to the nanoparticle backbone. The resulting silica nanoparticles can contain a large number of NIR dyes dependent on their size. For example some NIR fluorescent silica nanoparticle labels prepared that has an average radius around 15 nm, contains 16-20 covalently attached dye molecules inside of the nanoparticle. The primary applications of these particles are for bright fluorescent labels that can be used in applications such as immunochemistry, flow cytometry, and many other applications.

The fluorescent tracer agent 2,5-bis[N-(1-carboxy-2-hydroxy)]carbamoyl-3,6-diaminopyrazine, designated MB-102, has been developed with properties and attributes necessary for use as a direct measure of glomerular filtration rate (GFR). Comparison to known standard exogenous GFR agents in animal models has demonstrated an excellent correlation. A clinical trial to demonstrate this same correlation in humans is in progress. This clinical trial is the first in a series of trials necessary to obtain regulatory clearance from the FDA. We report herein the comparison of plasma pharmacokinetics between MB-102 and the known standard exogenous GFR agent Iohexol in healthy subjects with normal renal function. Post simultaneous administration of both agents, blood samples over a period of 12 hours were collected from each subject to assess pharmacokinetic parameters including GFR. Urine samples were collected over this same period to assess percent injected dose recovered in the urine. Results indicate MB-102 is a GFR agent in humans from the comparison to the standard agent.

Fluorescein (FL) and some of its precursors have proven to be effective fluorescent tracers in pharmaceutical and medical applications owing to their high quantum yield of fluorescence in physiological conditions and their high membrane permeability. In order to protect FL from metabolic effects during the process of its delivery, human serum albumin (HSA) has been used as a carrier because of its compatibility with the human body. In the present work, we used spectroscopic methods to characterize the binding mechanisms of FL and one of its derivatives, 5(6)- carboxyfluorescein (CFL), in the HSA protein. The absorbance change of the two ligands (FL and CFL) was quantified as a function of the HSA concentration and the results indicate a moderate binding strength for the two ligands inside HSA (1.00 ± 0.12 x 10⁴ M^-1). The quenching effect of FL(CFL) on the fluorescence intensity of W214 (the sole tryptophan in HSA) indicates that FL and CFL occupy Site I in the protein which is known to bind several hydrophobic drugs. By performing site-competitive experiments, the location of the ligands is determined to be similar to that of the anticoagulant drug warfarin. At higher ratios of [ligand]/[HSA], we observed an upward curvature in the Stern-Volmer plots which indicates that the ligands occupy more pockets in Site I, close to W214. Our results indicate that both ligands bind in HSA with a moderate strength that should not affect their release when used as fluorescent reporters. The chemical and physical identities of the two ligands are also preserved inside the HSA binding sites.

Fluorescent nanoparticles (NPs) such as KYb₂F₇:Tm³⁺ potential in biomedical applications due to their ability to absorb and emit within the biological window, where near infrared light is less attenuated by soft tissue. This results in less tissue damage and deeper tissue penetration making it a viable candidate in biological imaging. Another big factor in determining their ability to perform in a biological setting is the surface chemistry. Biocompatible coatings, including polyethylene glycol (PEG), polyvinylpyrrolidone (PVP), pluronic and folic acid are commonly used because they pose several advantages such as ease of functionalization, better dispersion, and higher cellular uptake. To study the effects of the NP surface chemistry, KYb₂F₇:Tm³⁺ a solvothermal method using PEG, PVP, pluronic acid, and folic acid as a capping agent, followed by thorough optical characterizations. Optical changes were thoroughly studied and compared using absorption, emission, and quantum yield data. Cell viability was obtained by treating Rhesus Monkey Retinal Endothelial cells (RhREC) with KYb₂F₇:Tm³⁺ and counting viable cells following a 24 hour uptake period. The work presented will compare the optical properties and toxicity dependency on the surface chemistry on KYb₂F₇:Tm³⁺. The results will also indicate that KYb₂F₇:Tm₃₊ nanoparticles are viable candidates for various biomedical applications.

Due to their robust optical properties, biological inertness, and readily adjustable surface chemistry, gold nanostructures have been demonstrated as contrast agents in a variety of biomedical imaging applications. One application is dynamic imaging of live cells using bioconjugated gold nanoparticles to monitor molecule trafficking mechanisms within cells; for instance, the regulatory pathway of epidermal growth factor receptor (EGFR) undergoing endocytosis. In this paper, we have demonstrated a method to track endocytosis of EGFR in MDA-MB-468 breast adenocarcinoma cells using bioconjugated gold nanocages (AuNCs) and multiphoton microscopy. Dynamic imaging was performed using a time series capture of 4 images every minute for one hour. Specific binding and internalization of the bioconjugated AuNCs was observed while the two control groups showed non-specific binding at fewer surface sites, leading to fewer bound AuNCs and no internalization.

Fluorescent tomography has been hindered by poor tissue penetration and weak signal which results in poor spatial resolution and quantification accuracy. Recently, it has been reported that activatable temperature responsive fluorescent probes which respond to focused ultrasound heating can improve the resolution and quantification of fluorescent tomography in deep tissue. This has lead to a new imaging modality, "Temperature-modulated fluorescent tomography." This technique relies on activatable thermo-sensitive fluorescent nanocapsules for whose fluorescence quantum efficiency is temperature dependent. Within a 4-5° C temperature range, the fluorescent signal increase more than 10-fold. In this molecular probe, Indocyanine Green (ICG) is encapsulated inside the core of a thermo-reversible pluronic micelle. Here we show the fluorescence response and temperature range of the nanocapsules which have been optimized for a higher temperature range to be used for in vivo animal imaging. We report on the feasibility of these temperature-sensitive reversible nanocapsules for in vivo applications by studying the pharmacokinetics in a subcutaneous mouse tumor model in vivo.

We developed a near infrared fluorophore-conjugated anti–Carcinoembryonic antigen (CEA) antibody for mice bearing tumor of CEA expressing cells, and demonstrated in vivo optical imaging by macro zoom microscopy. In the result, tumors of CEA-expressing cancer cells were specifically detected in vivo. Furthermore, cancer-specific fluorescence images were acquired at subcellular level in vivo by two-photon microscopy. In preclinical applications, the lymph node micrometastasis was also successfully visualized by two-photon microscopy. These results suggest that two-photon excitation microscopy in combination with an immunoconjugated probe could be widely adapted to cancer detection in clinical settings.

Water-driven self-assembly of lipids displays a variety of liquid crystalline phases that are crucial for membrane functions. In this work, we characterized the temperature-induced phase transitions in aqueous self-assembly systems using steady-state and time-resolved fluorescence measurements. The polar head group region was investigated using tryptophan (Trp) and two of its ester derivatives, and the hydrophobic tail region was probed using pyrene. The spectral changes in tryptophan and pyrene were used as a benchmark to estimate the polarity of the head group region and the tail region, respectively. A basic medium was detected and estimated for the polar region in the inverse cubic phase of a Guerbet glycolipid and was attributed to the structural effect of the narrow nanochannels. All the studied lipid compositions show completely reversible temperature-induced phase transitions, reflecting the thermodynamic equilibrium structures of their mesophases. The results reveal a large degree of heterogeneity and flexibility of the lipid self-assembly which may be crucial for carrying out different biological functions.

Oxidative stress plays a major role in many disease pathologies, notably in the central nervous system (CNS). For instance, after initial spinal cord injury, the injury site tends to increase during a secondary chemical injury process based on oxidative stress from necrotic cells and the inflammatory response. Prevention of this secondary chemical injury would represent a major advance in the treatment of people with spinal cord injuries. Few therapeutics are useful in combating such stress in the CNS due to side effects, low efficacy, or half-life. Mesoporous silica nanoparticles show promise for delivering therapeutics based on the formation of a porous network during synthesis. Ideally, they increase the circulation time of loaded therapeutics to increase the half-life while reducing overall concentrations to avoid side effects. The current study explored the use of silica nanoparticles for therapeutic delivery of anti-oxidants, in particular, the neutralization of acrolein which can lead to extensive tissue damage due to its ability to generate more and more copies of itself when it interacts with normal tissue.

Both an FDA-approved therapeutic, hydralazine, and natural product, epigallocatechin gallate, were explored as antioxidants for acrolein with nanoparticles for increased efficacy and stability in neuronal cell cultures. Not only were the nanoparticles explored in neuronal cells, but also in a co-cultured in vitro model with microglial cells to study potential immune responses to near-infrared (NIRF)-labeled nanoparticles and uptake. Studies included nanoparticle toxicity, uptake, and therapeutic response using fluorescence-based techniques with both dormant and activated immune microglia co-cultured with neuronal cells.

In recent years, infiltrating materials into the human body has become a great challenge many researches are facing. In medicine and cosmetics today, there are materials which are administrated to patients by injection only. The main challenge with topical medication is penetrating the skin barrier. The skin is an effective barrier between the body and the outside environment, which prevents foreign materials entering the body easily. However, reducing the size of the desired materials might help their skin penetration ability. Recently nanoparticles (NPs) are being evaluated for use in many fields like chemistry, biology, medicine, physics and optics. The technique used in this work for forming organic NPs (ONPs) is the application of sonic waves to an aqueous solution, known as sonochemistry. To investigate the physical penetration depth of ONPs into the human body, we first developed a novel optical technique for detecting NPs within tissues. The detection of NPs is done by the extraction and investigation of the reemitted light phase.

Finding new ways to deliver neurotrophic drugs to the brain in newborns is one of the contemporary problems of medicine and pharmaceutical industry. Modern researches in this field indicate the promising prospects of supramolecular transport systems for targeted drug delivery to the brain which can overcome the blood-brain barrier (BBB). Thus, the solution of this problem is actual not only for medicine, but also for society as a whole because it determines the health of future generations.

Phospholipid liposomes due to combination of lipo- and hydrophilic properties are considered as the main future objects in medicine for drug delivery through the BBB as well as increasing their bioavailability and toxicity. Liposomes functionalized by various proteins were used as transport systems for ease of liposomes use. Designing of modification oligosaccharide of liposomes surface is promising in the last decade because it enables the delivery of liposomes to specific receptor of human cells by selecting ligand and it is widely used in pharmacology for the treatment of several diseases.

The purpose of this work is creation of a coarse-grained model of bilayer of phospholipid liposomes, functionalized by specific to the structural elements of the BBB proteins, as well as prediction of the most favorable orientation and position of the molecules in the generated complex by methods of molecular docking for the formation of the structure. Investigation of activity of the ligand molecule to protein receptor of human cells by the methods of molecular dynamics was carried out.

Modern researches showed that nanocomposite films with magnetite nanoparticle incorporation have good perspectives for applications in electronics to create antireflective coatings and also for biomedical applications to create coatings with remote control of physical properties using alternative magnetic field or microwave radiation, which is very important for fabrication of new generation substrates in tissue engineering and advanced drug delivery systems. In particular, the unique properties of advanced nanocomposite microcapsules allowed developing of the supramolecular system of targeted drug delivery. A study of the behavior of the nanocomposite shell of microcapsules, which consists of alternate layers of negatively charged iron oxide nanoparticles and cationic polyallylamine hydrochloride molecules, was carried out.

The aim of the present study was to investigate the effect of the number of nanoparticle layers on magnetic properties of polyelectrolyte/nanoparticles nanocomposite microcapsules prepared via layer-by-layer technique using iron oxide colloids.

In result of numerical simulation using ANSYS Workbench software the behavior of the nanocomposite shell of microcapsules depending on the concentration of magnetite particles in it was investigated. Modal and harmonic analysis of behavior of the microcapsules shell was conducted in water at a temperature of 37^°. As a result of numerical experiment the eigenfrequencies and mode shape were first time defined for any modifications of the nanocomposite microcapsules. It has been established that the magnetic permeability value depends on the number of iron oxide nanoparticle layers in a nanocomposite microcapsule.

Model of terahertz emitter is based on carbon nanotube filled with fullerenes by the molecular-mechanical method. Creating of the conditions for which there was observed a terahertz range of was carried out. It is shown that the emitter has a nanometer size. It was established that the terahertz radiation with a frequency 0.6 THz is observed when there is no electric field, and this frequency can increase by an electric field of 1 * 10⁶ V / cm and the charge fullerene +5 e.

The complete search of controlling mode of motion of phospholipid was held. We found, that single molecule of phospholipid move randomly on graphene layer in both cases: with electric field or without it. And then we used additional charged molecules and changed value of electric field strength to find out how we can control that motion. We found, that motion of phospholipid molecule can be controlled through the using of molecules С₆₀ and electric field with strength 10⁶ V/m.

The investigation of energy stability of composites based on graphene sheets and carbon nanotubes was investigated. The dependence of enthalpy of formation from geometry parameters, such as length and diameter of nanotubes, distance between nanotubes on graphene sheet, of composites parts, optimal geometry for most energy sustainable composites were found.

At the present time carbon nanostructures are the main functional material for the development of electronic devices of broad applications. One of the most perspective for practical application forms of carbon nanostructures are nanotori. This paper presents results of prediction the stability of toroidal structures using computer modeling methods. The stability of carbon nanotori are determined by scanning of the local stresses map. Obtained results showed that the highest stability is characterized for carbon nanotori (13, 0) formed by the folding of a zigzag carbon nanotube. These nanotori are characterized by the lowest enthalpy of structure formation (101 eV).

In this paper we study the behavior of [2.2.2] cryptand with various metal ions (Na, K, Eu, Fe) inside the armchair carbon nanotube. To identify regularities of behavior for cryptand inside a nanotube we carried out a series of numerical experiments in molecular dynamics using Amber force field at different temperatures and under influence of external electric field. We have established the dependence of the oscillation frequency of the cryptand inside a carbon nanotube. On the basis of the established effect of fluctuations for cryptand inside the nanotube is predicted that such complexes can be used as miniature radiating systems.

Due to the increasing demand for functionalization of graphene and its application as a functional element of real electronic and / or mechanical devices, as well as due to its unique adhesive and sensory abilities the actual problem is the use of graphene as a substrate on which the assembly of supramolecular structures. Elements of such structures can be different molecules driven by external factors, and can be easily transported on graphene. These molecules primarily include miniature spheroidal fullerenes easy to navigate on the surface of graphene, in particular icosahedral C₆₀. The aim of this work was to find an effective method of manipulation of fullerene C₆₀ on graphene. As such method we proposed to introduce in graphene sheet structural defect of the atomic framework namely defect Stone-Wales (pentagon-heptagon pairs). Another structural defect studied in this paper is adsorbed on the Stone-Wales defect hydrogen atom. Molecular dynamics and tight binding method were applied to calculate the location of the molecule C₆₀ on graphene sheet and its movement. To identify the regulatities of behavior of fullerene on graphene sheet we carried out a series of numerical experiments at different temperatures. In this paper we calculated the energy profile of interaction between fullerene and graphene sheet. Obtained results showed that forming on the surface of the graphene sheet defects in a certain way, one can control the trajectory of molecules on graphene.

The main property of carbon nanotubes that determinates their wide application in electronics is a change of the chirality for ideal structure of a nanotube at implementing of structural Stone-Wales defect (pentagon-heptagon pairs) to its atomic framework. This property allows us to create nanotube-based different electronic devices (diodes, transistors, resistors), similar to traditional silicon devices. Nanotube with incorporated defect can be considered as a metalsemiconductor heterojunction. On the basis of this heterojunction semiconductor elements of very small size can be implemented, less than the current silicon elements. To create devices based on metal-semiconductor heterojunction is necessary to know the mechanisms of formation of the molecular compounds of nanotubes with different chirality. The aim of this work is a theoretical study of the formation of the molecular compounds of nanotubes with different chirality leading to the appearance of the metal-semiconductor heterojunction using molecular modeling methods. The object of investigation is a heterojunction formed by the compound of nanotubes with chirality indices (13,10) and (14, 10). To identify regularities of change in the electronic structure of the compound nanotubes we calculated the density of electronic states (DOS) for the heterojunction, and for each of its constituent chiral tubes. Also, we carried out a numerical evaluation of the reaction enthalpy of formation of the heterojunction. Based on these results it can be concluded that the investigated molecular compounds can be used to create highly sensitive sensors.

At the present time actual task of the modern materials is the creation of biodegradable biocompatible composite materials possessing high strength properties for medical purposes. One of the most promising biomaterials from a position of creation on their basis super strong nanofibres is chitosan. The aim of this work is a theoretical study of the structural features and physico-mechanical properties of biocomposite materials based on chitosan and carbon nanostructures. As matrix nanocomposite we considered various carbon nano-objects, namely carbon nanotubes and graphene. Using the developed original software complex KVAZAR we built atomistic and coarse-grained models of the biocomposite material. To identify regularities of influence of the configuration of the carbon matrix on the mechanical and electronic properties of biocomposite we carried out a series of numerical experiments using a classical algorithm of molecular dynamics and semi-empirical methods. The obtained results allow us to suggest that the generated biocomposite based on chitosan and carbon nanostructures has high stability and strength characteristics. Such materials can be used in biomedicine as a base material for creating of artificial limbs.

Many techniques in biological and clinical science use multiphoton absorbers for fluorescence. The applications include medical imaging for living cells, diagnostic techniques for disease and spectroscopy. The intrinsic value of the multiphoton absorber coefficients is therefore of the utmost importance. Additionally, the laser intensity at which the absorber saturates can determine which absorber, dye or protein is useful for a particular application.

Yet, experimental methods for determining the optical coefficients often yield different results. We describe several common methods of 2PA measurements and describe their features. As an example of the importance of applying the correct analysis to measurements, we fit experimental data and obtain values for multiphoton absorbers and accurately obtain their intrinsic values. Finally, we present the optical properties of several multiphoton materials used in biology.