Our ability to understand the structures and functions of living systems on a cellular and molecular level is mostly determined by the availability of imaging techniques capable of accessing a nanoscopic spatial resolution as well as providing structural information on molecular systems in vivo. While optical methods provide non-invasiveness, their spatial resolution is limited by a fundamental diffraction limit revealed more than 150 years ago by Ernst Abbe. This report addresses this grand-challenge and suggests a novel way of minimally invasive nanoscopic optical imaging inside a living cell. A powerful combination of optical tweezers, nonlinear optics and material science holds a promise of achieving unprecedented resolution of live-cell imaging, which should significantly advance our knowledge of molecular functions on a cellular level.

A photonic nanojet is a local field enhancement generated in the vicinity of a properly chosen microsphere or microcylinder illuminated by a collimated light beam. These photonic nanojets have waists smaller than the diffraction limit and propagate over several optical wavelengths without significant diffraction. We investigate the properties of photonic nanojets using rigorous solutions of Maxwell’s equations. A remarkable property we have found is that they can significantly enhance the backscattering of light by nanometer-scale particles (as small as ~1 nm) located within the jets. The enhancement factor for the backscattering intensity can be as high as five to six orders of magnitude. As a result, the observed intensity of the backscattered light from the dielectric microsphere can be substantially altered due to the presence of a nanoparticle within the light jet. Furthermore, the intensity and angular distribution of the backscattered signal is extremely sensitive to the size of the nanoparticle, which may enable differentiating particles with accuracy up to 1 nm. These properties of photonic nanojets make them an ideal tool for detecting, differentiating and sorting nanoparticles, which is of immense necessity for the field of nano-biotechnology. For example, they could yield potential novel ultramicroscopy techniques using visible light for detecting proteins, viral particles, and even single molecules; and monitoring molecular synthesis and aggregation processes of importance in many areas of biology, chemistry, material sciences, and tissue engineering.

We report on the development of a stroboscopic excitation technique using a self-pulsing laser, and show that it is a feasible method for obtaining fluorescence lifetime information from a biochip format. The self-pulsing lasers described here are versatile devices which have been used for one photon excitation measurements to determine the lifetime of cyanine 5 in water and ethanol. The same devices have been used to develop a technique whereby the emphasis for time-resolution of a lifetime measurement can be transferred to the excitation source from the detector and processing electronics by virtue of the multiple-pulse, variable frequency nature of the laser output.

Photonic crystals are emerging as an important class of engineered nanophotonic devices that possess unique optical properties and which can also provide textured surfaces for the study and control of cellular and molecular interactions. From among the many types of photonic crystal structures, two-dimensional (2D) and planar (slab) photonic crystals are the most attractive because of their ability to support guided-wave and active optical devices in semiconductor and polymer materials, serve as templates for device replication, and interface with colloidal and nanoparticle systems. This paper reports on the results of modeling and design efforts that show how 2d and slab silicon photonic crystals, based on their in-plane optical waveguiding and out-of-plane radiation properties, might be used to probe surface-bound cells and molecules or perform localized spectroscopy. The results of a parametric analysis show that photonic crystals comprised of high-index contrast materials (e.g. Si, air) are sensitive to geometric and material factors, potentially making them an effective medium to study molecular and cellular interactions critical to a number of biotechnological applications

Monte Carlo modeling was developed for simulating the light induced autofluorescence spectra of colon and cervical tissues at different dysplasia grades.These tissues were frozen sliced into 100 um thickness. The transmittance (Ta) and backscatter spectra (Ra) of these tissues were measured using double integral sphere system. The scattering coefficient, μs, and absorption coefficient, μa, were determined by the inverse adding-doubling (IAD) method from the Ta and Ra. spectra. The fluorescence intensity and spectra of these sliced tissues were measured using spectrometer system with cooled 2D CCD array. We simulated light energy distribution in tissue at 330 nm excitation and then convolution with fluorophores intensity and escape function in each tissue layer. The results of the simulation show: (1)fluorescence spectra change with different tissue characteristics, (2)fluorescence intensity decrease with the development of the dyplasia grades and the mucosa thickness, (3)the relative collagen signal decreases, hemoglobin signal increases, and NADH signal increases along with the dyplasia development. The simulated results matched well in vivo measured results. The approach provides an important means for understanding tissue fluorescence spectra’s changes that are very critical for clinic diagnosis.

One of the major goals of biosensor technology is to detect and quantify in detail analytes with very high accuracy. To achieve this, much of the emphasis in sensor fabrication has been laid on antibody-antigen interaction. The consequence of this focus of enzyme biosensor studies is the development of critical techniques which can be extended in the detection of Acute Myocardial Infarction (AMI). Biosensors for AMI have attracted considerable interest in the last few years since the monitoring of a specific substance is central in enzymatic reactions. This interest has led to the investigation of biochemical markers of myocardial injury. These biomarkers facilitate the diagnosis and treatment of patients with AMI. Serial measurements of biochemical markers are now universally accepted as an important determinant in AMI diagnosis. Due to their high sensitivity and specificity over other biomarkers, the troponins are the markers of choice for the diagnosis or exclusion of AMI. The present techniques used in the identification of the troponins are lengthy and require large amount of specimen solution. The present research is directed towards the identification of optical detection procedures that are compatible to the miniaturization. In the present study an effort has been made to study the antigen-antibody reaction of rabbit skeletal muscle troponin C (TnC) and bee venom melittin (ME). Fluorescence energy transfer experiments were done to investigate the Ca ²⁺ -dependant interaction of TnC-ME in a 1:1 complex. Experiments were also conducted on TnC-ME binding at different ratios. These results validate the biosensor technology and illustrate how a biosensor can be developed based on the study of interaction between monoclonal antibody and antigen reaction in real time. The reported experimental results provide valuable information that will be useful in the development of a biosensor for the detection of AMI.

Silicon quantum dots have been synthesized in micelles. Particle sizes have been ascertained by transmission electron microscopy and UV-Vis absorption and photoluminescence spectroscopy. The surface of the silicon particles produced have been modified to produce hydrophobic and hydrophilic particles by reaction with either with 1-heptene or allylamine respectively. For biological applications control of the surface character of the nanocrystals is essential. FTIR spectra show the surface modification of the particles by 1-heptene or allylamine.

Fluorescent semiconductor nanoparticles, or quantum dots, have potential uses as an optical material, in which the optoelectronic properties can be tuned precisely by particle size. Advances in chemical synthesis have led to improvements in size and shape control, cost, and safety. A limiting step in large-scale production is identified to be the raw materials cost, in which a common synthesis solvent, octadecene, accounts for most of the materials cost in a batch of CdSe quantum dots. Thus, less expensive solvents are needed. In this paper, we identify heat transfer fluids, a class of organic liquids commonly used in chemical process industries to transport heat between unit operations, as alternative solvents for quantum dot synthesis. We specifically show that two heat transfer fluids can be used successfully in the synthesis of CdSe quantum dots with uniform particle sizes. We observe differences in particle growth using the various solvents.

InP quantum dots (QDs) with zinc blende structure and InN QDs with hexagonal structure were synthesized from appropriate organometallic precursors in a noncoordinating solvent using myristic acid as a ligand. The QDs were characterized by TEM, the associated energy dispersive spectroscopy (EDS), electron diffraction, and steady state UV-VIS optical absorption and photoluminescence spectroscopy. To our best knowledge, this paper reports synthesis of InN colloidal quantum dots for the first time.

The present paper introduces to the problems related with the application of silica shells to luminescent semiconductor nanocrystals, especially for bioanalytical applications. Two examples for the preparation of silica shells are presented: First, preparation of a very thin silica layer with simultaneous functionalisation and biological application. Second, a new preparation method for silica shells based on a sol-gel approach. In summary, it is shown, that --- despite of all problems --- high-quality silica coated nanocrystals can be prepared and are well suited for bioanalytical applications.

We have developed supercritical hydrothermal synthesis method of nanoparticles. In the method, metal salt aqueous solution is mixed with high temperature water to rapidly increase the temperature of the metal salt solution and thus reduce the reactions and crystallizations during the heating up period. By using this method, we succeeded in the continuous and rapid production of nanocrystals. In this paper, we propose a new method to synthesize organic-inorganic fused materials based on the methods of supercritical hydrothermal synthesis. By introducing organic materials in a reaction atmosphere of supercritical hydrothermal synthesis, nanoparticles whose surface was modified with organic materials were synthesized. In supercritical state, water and organic materials form a homogeneous phase, which provides an excellent reaction atmosphere for the organic modification of nanoparticles. Modification with bio-materials including amino acids was also possible. By changing organic modifiers, particle morphology and crystal structure were changed. This organic surface modification provides a various unique characteristics for the nanoparticles: Dispersion of nanoparticles in aqueous solutions, organic solvents or in liquid polymers can be controlled by selecting hydrophilic or hydrophobic modifiers. Polymer-like materials can be formed for the amino acid modified nanoparticles probably by the self-assembly of amino acid.

We report a simple and rapid method to synthesize water-soluble and biocompatible fluorescent quantum dot (QD)-micelles by encapsulation of monodisperse, hydrophobic QDs inside surfactant/lipid micelles. Analysis of UV-vis spectra, transmission electron microscopy, and photo luminescence spectra indicate that the water-soluble semiconductor QD-micelles are monodisperse and retain the optical properties of the original hydrophobic QDs. The QD-micelles were shown to be biocompatible and exhibited little or no aggregation when taken up by cultured rat hippocampal neurons.

The longer the excitation light is dosed on the quantum dots, the larger the intensity of the fluorescent light we get. It is known as the light memory effect. This effect was found in dry film of quantum dot at first. We found this memory effect also occurs inside the cell and also found the wavelength of the fluorescent light to shift toward shorter. In order to acquire the water solubility, we use mercapto undecanic acids, mercapto glycerol, and mercapto amine for the surface treatment. The base line of the fluorescent intensity is the highest with the quantum dot treated with mercapto undecanoic acids. That with the mercapto glycerol is the next. That with the mercapto amine is the lowest among the three compounds. We measured the zeta potential of the surface of these three different treated quantum dot. The quantum dot treated with the mercapto undecanoic acids is negatively charged, that with the mercapto amine is positively charged and that with the mercapto glycerol is the neutral. We added the electron donor components and the electron scavenger components and found that the electron donor components raises up the intensity of the fluorescent light and the electron scavenger components puts down the intensity, as expected. The conjugation of the quantum dot with the bio-molecule such as protein, sugar and nucleic acids will change the zeta potential which lead to speculate that the measurement of the zeta potential of the bio-molecule could predict roughly the fluorescent intensity of the conjugated bio-molecule and quantum dot complex.

Transposable elements (TEs) or transposons are mobile segments of DNA that are capable of being excised and moved from one chromosomal location to another by a process known as transposition. This process requires an enzyme called the transposase that performs the excision reaction, recognizes specific target site sequences and then promotes insertion of the TE at the target site (transposition). This study provides new clues towards unraveling the causes behind the preferential affinity of the Hermes transposable element for certain insertion sites compared to other sequences which also contain recognizable target sites. The technique consists of a rapid, simple and reproducible assay that can be used to detect differences in the ability of various oligonucleotides to influence the aggregation of colloidal gold nanoparticles. The aggregation of the gold nanoparticles is monitored through UV-Visible absorption spectroscopy. Single isolated colloidal gold particles have a surface plasmon resonance manifested as a single absorbance peak at approximately 520 nm and aggregated gold complexes develop new red-shifted peaks/shoulders depending on the nature and extent of the aggregated complex. A simple ratiometric study of the area under the single and aggregated plasmon resonance peaks gives information about the extent of the aggregation. It is postulated that differences in dynamic flexibility of the oligonucleotides affect their influence on the aggregation state of the gold nanoparticles. Therefore such differences in dynamic flexibility between various insertion sites could directly or indirectly contribute to the observed target site preferences of the Hermes transposable element.

Detection tags based upon surface enhanced Raman scattering provide an alternative to the widely used fluorescence methods. Several aspects of these tags are presented in this report. The tags can be made to display many different spectra, thus they can be used for multiplexed detection schemes. They generate a large enough number of photons to be readily detected, and spectra acquired from mixtures of tags can be analyzed giving accurate amounts of the components. The surface of the tags can be easily modified to present common biological molecules (streptavidin and analogues). Finally, we demonstrate their use to quantitatively detect interleukin-4 (IL4) and interleukin-7 (IL7) in a microarray format.

An electron transfer pathway between Cytochrome c (Cyt c) molecules, a 30nm Au nanoparticle and an ITO working electrode in an electrochemical cell is constituted by molecular junctions. The scattering spectrum of single Au nanoparticle is measured simultaneously with the cyclic voltammogram. The plasmon resonance wavelength and the scattering cross section of the single Au nanoparticle are affected by the redox reactions of less than 200 cyt c molecules on its surface and exhibit cyclic variations. The electron shuttling between Cyt c molecules and ITO electrode through the Au nanoparticle in the redox process as well as the conformation change of the Cyt c molecules between ferric and ferrous states are accounted the reasons for the change of the plasmon resonance wavelength and scattering cross section. The presented study of the interaction of biological electron transporter protein and photonic nanostructure provide a nano-scale system to probe the electron transfer events in the biological system. It also has the implicational importance to the development of future hybrid bio-optoelectronic devices.

We describe a versatile scheme to prepare an array of multidentate surface capping molecules. Such materials permit strong interactions with semiconductor nanocrystals and render them water compatible. These ligands were synthesized by reacting various chain length poly (ethylene glycols) with thioctic acid, followed by ring opening of the dithiolane moiety. Functionalization of CdSe-ZnS quantum dots with these ligands allow processing of the nanocrystals not only in aqueous but in many other polar solvents. Further synthetic processing of the ligands with biotin moieties allowed for investigating assays based on the avidin-biotin interactions. These ligands provide a straightforward means of preparing QDs that exhibit greater resistance to environmental changes, making them more amenable for use in live cell imaging and other biotechnological applications.

Colloidal NCs consist of an inorganic particle and an organic coating that determines their solubility, functionality, and influences their photophysics. In order for these NCs to be biocompatible, they must be water-soluble, nontoxic to the cell, and offer conjugation chemistries for attaching recognition molecules to their surfaces. In addition they should efficiently target to biomolecules of interest, be chemically stable, and preserve their high photostability. The requirements for their application in single-molecule biological studies are even more stringent: fluorescent NCs should be monodisperse, have relatively small size (to limit steric hindrance), reduced blinking, large saturation intensity, and high quantum yield (QY).

Water-soluble quantum dots (qdots) have been introduced as bright fluorophores into life sciences research. Although various photophysical pathologies of qdots have been found, how their biological applications will be affected --- particularly in the native biological environment --- has not been evaluated. By fluorescence coincidence analysis and fluorescence cross-correlation spectroscopy, we studied the dark fraction of free-diffusing qdots in aqueous solution. We were able to detect individual qdots and found significant heterogeneity --- well-distinguished dark qdots and bright qdots. We estimated the bright fraction of Qdot525-Streptavidin to be about 55%. Blinking events were also noticed in fluorescence coincidence analysis, with “on/off” timescale from submilliseconds to tens of milliseconds.

Quantum dots have opened up a plethora of possibilities in biological detection and imaging. Their small size, stable luminescence and resistance to photobleaching make them ideal for intracellular imaging and detection. It is highly desirable to develop tools that will allow detection and imaging of biological processes without disrupting native cellular processes. A significant barrier to use of quantum dots in living cells is ability to deliver and detect single quantum dots inside living cells. In this article we describe a bacterial toxin dependent method that allows delivery of single quantum dots. We also demonstrate use of a single molecule detection system to detect both single and aggregated quantum dots in living cells in real time. We compare results of quantum dot delivery from receptor mediated endocytosis and HIV-TAT peptide mediated delivery methods with the bacterial toxin Streptolysin O. Our results show that Streptolysin O is able to deliver single quantum dots to living cells. Our results also indicate that the mechanism of cargo delivery by HIV-TAT peptide might be endocytosis dependent. Ongoing work in this direction involves showing that single, functional quantum dot probes can also be delivered using the bacterial toxin.

Cancer of the prostate affects approximately 1 in 11 men. Current early screening for prostate cancer utilizes digital rectal examinations to detect anomalies in the prostate gland and blood test screenings for upregulated levels of prostate specific antigen (PSA). Many of these tests are invasive and can often be inconclusive as PSA levels may be heightened due to benign factors. Prostate specific membrane antigen (PSMA), a well-characterized integral membrane protein, is expressed in virtually all prostate cancers and often correlates with cancer aggressiveness. Therefore, it may be used as an indicator of cancer growth and metastases. PSMA-specific antibodies have been identified and conjugated to fluorescent markers for cancer cell targeting; however, both the antibodies and markers possess significant limitations in their pharmaceutical and diagnostic value. Here we report the use of semiconductor nanocrystals bioconjugated to PSMA-specific aptamer recognition molecules for prostate carcinoma cell targeting. The nanocrystal/aptamer bioconjugates are small biocompatible probes with the potential for color-tunability for multicolor imaging. Ongoing in vitro and in vivo research seeks to introduce these nanoparticle bioconjugates into medical diagnostics.

Quantum dots (QDs) are a versatile synthetic photoluminescent nanomaterial whose chemical and photo-physical properties suggest that they may be superior to conventional organic fluorophores for a variety of biosensing applications. We have previously investigated QD-fluorescence resonance energy transfer (FRET) interactions by using the E. coli bacterial periplasmic binding protein - maltose binding protein (MBP) which was site-specifically dye-labeled and self assembled onto the QD surface and allowed us to monitor FRET between the QD donor and the acceptor dye. FRET efficiency increased as a function of the number of dye-acceptor moieties arrayed around the QD donor. We used this system to further demonstrate a prototype FRET based biosensor that functioned in the chemical/nutrient sensing of maltose. There are a number of potential benefits to using this type of QD-FRET based biosensing strategy. The protein attached to the QDs surface functions as a biosensing and biorecognition element in this configuration while the QD acts as both nanoscaffold and FRET energy donor. In this report, we show that the sensor design can be extended to target a completely unrelated analyte, namely the explosive TNT. The sensor consists of anti-TNT antibody fragments self-assembled onto the QD surface with a dye-labeled analog of TNT (TNB coupled to AlexaFluor 555 dye) prebound in the fragment binding site. The close proximity of dye to QD establishes a baseline level of FRET and addition of TNT displaces the TNB-dye analog, recovering QD photoluminescence in a concentration dependent manner. Potential benefits of this QD sensing strategy are discussed.

We have developed conjugates with quantum-dots (QDs) for the purpose of analysis of nanosystems that are organic or inorganic in nature such as DNA and carbon nanotubes. First, by employing Florescence Resonant Energy Transfer (FRET) principles, a hybrid molecular beacon conjugates are synthesized. For water- solubilization of QDs, we modified the surface of CdSe-ZnS core-shell QD by using mercaptoacetic acid ligand. This modification does not affect the size of QDs from that of unmodified QDs. After linking molecular beacons to the carboxyl groups of the modified QDs using 1-Ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride, hybrid molecular beacons are prepared as a DNA probe. After hybridization with specific target DNA and non-specific target DNA, the hybrid conjugates show high specificity to the target DNA with 5-fold increase in the intensity of fluorescence. By developing atomic model of the conjugates, we calculated with 8 numbers of molecular beacons on a single quantum dots, we could increase the efficiency of FRET up to 90%. In other hands, for application of quantum dots to the carbon nanotubes, FRET is a barrier. Thus, after employing 1 % sodium-dodecyl-sulfonate (SDS), single-walled carbon nanotubes are decorated with QDs at their outer surface. This enables fluorescent microscopy imaging of single-walled carbon nanotubes which is a more common technique than electron microscopy. In summary, QDs can be used for analysis or detection of both organic and inorganic based nanosystems.

Steady state and time-resolved fluorescence measurements were used to investigate the ability of luminescent quantum dots (QDs) to function as efficient energy acceptors in fluorescence resonance energy transfer (FRET) binding assays with organic dye donors. Fluorescent dyes, AlexaFluor 488 or Cy3, were used with various QD acceptors in QD-dye-labeled-protein conjugates. Data derived from both sets of experiments showed no apparent FRET from dye to QD. The collected data were discussed within the framework of a competition between a fast radiative decay rate of the donor excitation and a slower FRET decay rate. This is due to the long exciton lifetime of the acceptor compared to that of the dye, combined with substantial QD direct excitation.

Lanthanide-ion doped oxide nanoparticles were functionalized for use as fluorescent biological labels. These nanoparticles are synthesized directly in water which facilitates their functionalization, and are very photostable without emission intermittency. Nanoparticles functionalized with guanidinium groups act as artificial toxins and specifically target sodium channels. They are individually detectable in cardiac myocytes, revealing a heterogeneous distribution of sodium channels. Functionalized oxide nanoparticles appear as a novel tool particularly well adapted to long-term single-molecule tracking.

Fluorescence imaging is a commonly used approach for sensitive, non-invasive, high resolution imaging of live cells and organisms. The full potential of this approach, however, has not been realized due to restrictions imposed by routinely used organic fluorophores. Use of inorganic fluorescent nanocrystals, quantum dots (QDs), provides an opportunity to better realize the potentials of live cell fluorescent imaging. A prerequisite to the use of QDs for cellular and biomedical imaging, however, is developing approaches for labeling live cells with these reporters and establishing the safety of these reporters for imaging live cells and organisms. Here we present a variety of approaches to efficiently tag molecules and live cells with QDs and have used these methods for the long-term tracking of multiple populations of cells during growth and development. Using these approaches we demonstrate that QD-labeling has no detectable effects on either the physiology of the cells or the organism in which they reside. This establishes QD as a suitable fluorescent probe for both live cells and in vivo imaging.

The interface of targeting molecules that can recognize and identify specific biomolecules with highly luminescent semiconductor nanocrystals or quantum dots can lead to a novel and powerful new class of probes for studying biomolecules in real-time or for imaging and detecting diseases. We describe the rationale design of optical nanoprobes by using fluorescent semiconductor quantum dots with targeting molecules (TMs)-identified using phage display screening. Quantum dots are nanometer-sized particles with unique and tunable optical properties. They offer numerous optical advantages over traditional organic fluorophores in biological analysis and detection (e.g., photostability, continuous absorption profile).

Quantum dot bioconjugates offer unprecedented opportunities for monitoring biological processes and molecular interactions in cells, tissues, and organs. We are interested in developing applications that permit investigation of physiological processes and cytoskeletal organization in live cells, and allow imaging of complex organs, such as the auditory and vestibular sensory structures of the inner ear. Multiphoton microscopy is a powerful technique for acquiring images from deep within a sample while reducing phototoxic effects of laser light exposure on cells. Previous studies have established that a solid-state Nd:YLF laser can be used to acquire two-photon and three-photon images from live cells while minimizing phototoxic side effects (Wokosin et al., 1996, Bioimaging, 4:208-214; Squirrell et al., 1999, Nature Biotechnology, 8:763-767). We present here the results of experiments using an all-solid-state Nd:YLF 1047 nm femtosecond laser (Microlase DPM1000) source to excite quantum dot bioconjugates. Cells were labeled with Qdot (Quantum Dot Corporation) bioconjugates or with Alexa Fluor (Molecular Probes) bioconjugates and then imaged with a BioRad 1024 confocal microscope configured for multiphoton imaging using internal or external (non-descanned) detectors. Results demonstrate that the Nd:YLF laser can be used to stimulate fluorescence emission of quantum dots and Alexa Fluor bioconjugates in cultured amphibian (Xenopus) and mammalian (rat, chinese hamster) cells. We conclude that the Nd:YLF laser is a viable excitation source that extends the applicability of quantum dots for investigation of biological processes using multiphoton microscopy.

The use of semiconductor quantum dots (qdots) as optical imaging agents is well established; however, the use of these nanoparticles to form interfaces that interact directly with cells has proved more challenging. Our goal has been to create specific nanoparticle-neuronal receptor interfaces that can be optically excited to elicit an action potential. Our preliminary calculations showed the possible feasibility of this approach, even in the presence of Debye screening. We therefore have developed several methods of nanoparticle-directed binding to cell surfaces. These methods can produce either nonspecific or directed interfaces, only nanometers from the receptor of interest. These techniques are versatile and have allowed us to achieve a variety of nanoparticle-neuron interfaces; however, they are also subject to inherent limitations at the cell surface, including endocytosis. To address this concern, we have developed tethered nanoparticle films, which may be able to interact nonspecifically with receptors of nerve cells cultured on their surfaces. These films were found to be extremely stable in cell culture media, but degraded within 3-5 days in primary neuron cultures. Here, we discuss the initial development of tethered quantum dot films produced in our laboratory and their compatibility with cell culture conditions.

Photodynamic therapy (PDT) is an emerging therapy for cancer treatment that shows the greater selectivity towards the malignant cells. Semiconductor nanoparticles are a novel class of photosensitizers with properties that are not easily available with conventional PDT reagents. Their potential properties such as improved luminescence, resistance to photobleaching, and the possibility to modify the surface chemically make them suitable candidates for PDT. In this report, we discuss the synthesis of ternary CdSe_1-x Tex nanoparticles along with well known CdSe QDs and their potential in generating the singlet oxygen state by Foerster Resonance Energy Transfer (FRET) to a PDT reagent or by direct triplet-triplet energy transfer to molecular oxygen.

The fluorescence intermittency of single, bare, CdSe/ZnS quantum dots was probed using single molecule confocal microscopy and found to demonstrate power law kinetics. Various threshold values and line fitting parameters are employed in the data analysis and their effects on the extracted power law exponents, m_off and m_on, are presented. The threshold is found to be critical in determining m_off while having no significant effect on m_on. The mean plus 2σ threshold, calculated from the background noise in the measurement, results in a more negative m_off slope in comparison to the mean plus 3σ or mean plus 4σ thresholds. This is likely due to the mean plus 2σ threshold lying within the background noise outliers which mimic short on events. In contrast, the mean plus 4σ threshold is above 99.99% of the background noise while adequately below the fluorescence signal. Additionally, it is found that fitting only the ten most probable data points rather than all the data points in the log-log probability density graphs results in no significant change in m_off and mon.

Fluorescent nanoparticles, such as nanocrystal quantum dots (QDs), novel nanometer-size probes and have the potential to be used as easy imaging tool for molecular biology and bioimaging including medical applications, since some nanocrystals emit higher and far longer fluorescence than conventional organic probes. QDs are now becoming widely used in biotechnology and medical applications. QDs have several advantages over organic fluorophores with regard to high luminescence, stability against photobleaching, and a range of fluorescence wavelengths from blue to infrared depending on the particle size. In this review, we reported labeling of some kinds of immune cells and biomolecules with several QDs coated with hydrophilic carboxyl/amine groups, and reported that we could image the circulation of mouse lymphocytes in vivo by QDs. In addition, we also reported here about the cytotoxicity of these nanocrystals.

Photo-luminescent semiconductor quantum dots are nanometer-size probes that have the potential to be applied to the fields of the bio-imaging and the study of the cell mobility inside the body. At the same time, on the other hand, quantum dots are expected to carry some kind of molecules to the local organ inside of the animal body, which leads to the expectation that they can be used as a medicine-carrier. For this purpose, we conjugate (2S)-1-[(2s)-2-Methyl-3-sulfanylpropionyl]pyrrolidine-2-carboxylic acid (cap) with the quantum dot. Cap has the effect as an anti-hypertension drug, which inhibits angiotensin 1 converting enzyme. We conjugated the quantum dot with cap by the exchange reaction avoiding the regions which holds medicinal effect. Quantum dot conjugated with cap (QD-cap) were 3-times brighter than thioglycerol-coated quantum dots (QD-OH). The particle size of cap was 1.1nm and that of QD-cap was 12nm. QD-cap was permeated into the HeLa cells, while QD-MUA were taken into the HeLa cells by endocytosis. In addition, no apoptosis was detected against the cells that permeated QD-cap, because there was no damage to DNA. These results indicated that QD-conjugated medicines (QD-medicine) could be safe in the experiment on the level of the cell. More over, when QD-cap was intravenously injected into Stroke-prone Spontaneously Hypertensive Rats (SHRSP), they reduced blood pressure at systole. Therefore, the anti-hypertension effect of cap remained after conjugated with the quantum dot. These results suggested that QD-medicine were effective on the animal level.

Here we show that femtosecond laser irradiation can be used to evoke dynamic calcium concentration changes in living cells. The relatively localized interaction that results from the two-photon absorption process allows the release of calcium from intracellular stores in cells in vitro. The self-catalytic response to calcium elevation in a cell can increase the initial release of calcium further so that the entire cell undergoes a rise in cytosolic calcium concentration (i.e. a calcium wave). The calcium stimulation was observed in HeLa (non-excitable) and PC12 (excitable) cells, and could be seen to occur inside a range of power levels between approximately 20 to 80mW. The observation of direct calcium release by femtosecond laser which leads to a calcium wave in the cell has implications for photolytic calcium uncaging experiments since it could be a competing, or even dominant factor in some experiments using caged calcium for the generation of calcium waves.

The surface coating of quantum dots has been characterised using Z-stem. Quantum dots have been pegylated to increase stability in aqueous solution. The fluorescence intensity of the quantum dots was modulated pegylation. PEG was coupled using different ratios of EDC, PEG and NHS. Optimum coupling conditions were found to occur when 2000 equivalents of PEG were reacted with 1 equivalent of dot in the presence of 1500 equivalents of NHS and EDC. Angiotensin II was also conjugated to quantum dots and these conjugates were shown to be biologically active. Quantum dots have also been surface functionalised with other peptides such as NGR with subsequent demonstration of cell surface binding and can be characterized by flow cytometry.