This PDF file contains the front matter associated with SPIE Proceedings Volume 8212, including the Title Page, Copyright information, Table of Contents, and the Conference Committee listing.

The ease and flexibility of functionalization and inherent light scattering properties of plasmonic nanoparticles make them suitable contrast agents for measurement of cell surface markers. Immunophenotyping of lymphoproliferative disorders is traditionally undertaken using fluorescence detection methods which have a number of limitations. Herein, surface-enhanced Raman scattering (SERS) gold nanoparticles conjugated to monoclonal antibodies are used for the selective targeting of CD molecules on the surface of chronic lymphocytic leukemia (CLL) cells. Raman-active reporters were physisorbed on to the surface of 60 nm spherical Au nanoparticles, the particles were coated with 5kDa polyethylene glycol (PEG) including functionalities for conjugation to monoclonal IgG1 antibodies. A novel method for quantifying the number of antibodies bound to SERS probes on an individual basis as opposed to obtaining averages from solution was demonstrated using metal dots in transmission electron microscopy (TEM). The specificity of the interaction between SERS probes and surface CD molecules of CLL cells was assessed using Raman spectroscopy and dark field microscopy. An in-depth study of SERS probe targeting to B lymphocyte marker CD20 was undertaken, and proof-of-concept targeting using different SERS nanoparticle dyes specific for cell surface CD19, CD45 and CD5 demonstrated using SERS spectroscopy.

60 nm diameter gold nanoparticles (AuNP) were coated with a ternary mixture of lipids and targeted to human lymphocytes. Previously, the versatility, stability and ease of application of the lipid coating was demonstrated by the incorporation of three classes of Raman-active species. In the present study, lipid encapsulated AuNPs were conjugated to two targeting species, namely whole antibodies and antibody fragments (Fab), by two methods. Furthermore, in vitro targeting of lipid-encapsulated Au nanoparticles to patient-derived chronic lymphocytic leukemia (CLL) cells was demonstrated by Raman spectroscopy, Raman mapping, and darkfield microscopy. These results further demonstrate the versatility of the lipid layer for imparting stability, SERS activity, and targeting capability, which make these particles promising candidates for biodiagnostics.

The modeling, fabrication and characterization of PSi fabricated from both (110) and (100) surface oriented silicon for optical sensing is thoroughly reported. First, based on the generalized Bruggeman method, the birefringence and sensitivity of the fabricated membranes were calculated as a function of the fabrication parameters such as porosity and pore sizes; and external effects, such as the pores surface oxidation. Thereafter we report on the fabrication of PSi membranes from (110) and (100) surface oriented silicon with pore sizes in the range of 50 - 80 nm, and the characterization of their birefringence using a polarimetric setup. Their sensitivities were determined by filling the pores with several liquids having different refractive index. As a result, sensitivities as high as 1407 nm/RIU were obtained for the (110) samples at a 1500 nm wavelength and 382 nm/RIU for the (100) samples at the same wavelength.

While Q ~ 1million has been demonstrated in freely suspended photonic crystal (PC) membranes, the reduced refractive index contrast when PC microcavities are immersed in phosphate buffered saline (PBS), a typical ambient for biomolecules, reduces Q by more than 2 orders of magnitude. We experimentally demonstrate photonic crystal microcavity based resonant sensors coupled to photonic crystal waveguides in silicon on insulator for chemical and bio-sensing. Linear L-type microcavities are considered. In contrast to cavities with small modes volumes but low quality factors for bio-sensing, we show that increasing the length of the microcavity enhances the quality factor of the resonance by an order of magnitude and also increases the resonance wavelength shift while still retaining compact device characteristics. Q~26,760 and sensitivity down to 7.5ng/ml and ~9pg/mm² in bio-sensing was experimentally demonstrated in SOI devices for goat anti-rabbit IgG antibodies with K_d~10^-6M. The increase in cavity length follows from fundamental engineering limitations in ink-jet printing or microfluidic channels when unique receptor biomolecules are coated on separate adjacent sensors in a microarray.

Cavity ring down measurement approach is a promising technique for biosensing as it is insensitive to intensity uctuations of a laser source. This technique in conjunction with ultra high Q microcavities have a great potential for ultra sensitive biosensing. Until now, most work on microcavity biosensors has been based on measurement of the resonant frequency shift induced by binding event on surface of the microcavity. Such measurements suer from the noise due to intensity uctuations of the laser source. However, the binding event will also introduce shift in quality factor of the microcavity, which can be tracked by using cavity ring down spectroscopy. In this work, we report on experimental demonstration of application of ring down measurement approach to microcavities for biosensing by tracking disassociation phase of a biotin-streptavidin reaction. These measurements were performed by using a bioconjugated ultra high Q microtoroidal cavity immersed in a liquid microacquarium. We found that disassociation curves agree with previously reported results on the protein kinetics measurements.

Optofluidics organically integrates microfluidics and photonics and is an emerging technology in biological and chemical analysis. In this paper, we overview the recent studies in bio-chemical sensing applications of optofluidics. Particularly, we report the research progress in our lab in developing diverse optofluidic devices using two unique configurations: thin-walled capillary based optofluidic ring resonator (OFRR) and multi-hole capillary based optofluidic platforms. The first one has been developed to be OFRR-based label-free biosensor, microfluidic laser based intra-cavity sensors, and on-column optical detectors for micro-gas chromatography (μGC), while the second one has been developed to be optofluidic Fabry-Pérot based label-free biosensor and optofluidic Surface-Enhanced Raman Spectroscopy (SERS) biosensor. All of these devices take advantage of superior fluidic handling capability and high sensitivity, and have been used in detecting various biological and chemical analytes in either liquid or vapor phase.

Optoelectronic tweezers (OET) allows the spatial patterning of electric fields through selected illumination of a photoconductive surface. This enables the manipulation of micro particles and cells by creating non-uniform electrical fields that then produce dielectrophoretic (DEP) forces. The DEP responses of cells differ and can produce negative or positive (repelled or attracted to areas of high electric field) forces. Therefore OET can be used to manipulate individual cells and separate different cell types from each other. Thus OET has many applications for medical diagnostics, demonstrated here with work towards diagnosing Human African Trypanosomiasis, also known as sleeping sickness.

Surface-enhanced Raman scattering (SERS) has emerged as a powerful analytical technique for direct detection of chemical and biological analytes because of high sensitivity, selectivity, and rapid response. Here we propose and develop a novel optofluidic SERS structure, i.e., nanoparticle-functionalized flow-through multihole capillary. This unique platform provides many advantages. First, its 3-dimensional (3-D) structure, similar to nanoporous aluminum membranes, nanoporous polymer monoliths, and photonic crystal fibers (PCFs), provides large surface area for the deposition of noble nanoparticles or nanoclusters to achieve high SERS intensity. Second, it has well-defined flow-through channels. It provides robust and controllable nanoparticle immobilization like PCFs, but much higher nanoparticle density thus large SERS-active sites due to large surface within the detection volume, and also enables fast and convenient analyte delivery for real-time, online detection. Third, the well-defined multihole capillary can also confine and transmit light along the longitudinal direction, accumulating large SERS signal like PCFs. Fourth, it is easy to integrate with other sensing platforms, such as label-free biosensors, to provide comprehensive information on molecular interaction. Moreover, the multihole capillary can be mass-produced easily and cost effectively using the fiber drawing method. In this report, using a capillary consisting of thousands of micrometer-sized holes adsorbed with gold nanoparticles, we investigated the proposed optofluidic SERS system using the transverse and longitudinal detection methods, where the SERS excitation and collection were perpendicular to and along the capillary, respectively. A detection limit better than 100 fM for rhodamine 6G was achieved with an enhancement factor exceeding 10⁸.

Analysis of the intrinsic scatter and fluorescence profiles of marine algae can be used for general classification of organisms based on cell size and fluorescence properties. We describe the design and fabrication of a Microflow Cytometer on a chip for characterization of phytoplankton. The Microflow Cytometer measured distinct side-scatter and fluorescence properties of Synechococcus sp., Nitzschia d., and Thalassiosira p. Measurements were confirmed using the benchtop Accuri C6 flow cytometer. The Microflow Cytometer proved sensitive enough to detect and characterize picoplankton with diameter approximately 1 mm and larger phytoplankton of up to 80 mm in length. The wide range in size discrimination coupled with detection of intrinsic fluorescent pigments suggests that this Microflow Cytometer will be able to distinguish different populations of phytoplankton on unmanned underwater vehicles. Reversing the orientation of the grooves in the channel walls returns the sample stream to its original unsheathed position allowing separation of the sample stream from the sheath streams and the recycling of the sheath fluid.

This paper reports a micro flow cytometer fabricated in Polymethylmethacrylate (PMMA) in which a 3d hydrodynamic flow focusing is employed in order to align the particles in a single line along the focused stream. The device has been fabricated by direct micro milling of two parts of PMMA that were finally bonded together. With a suitable choice of the fluidic channel geometry, a circular sample stream located in the center of the channel is obtained. Numerical simulations have been performed in order to investigate the flow characteristic of the structure and find the desiderated geometry. Three dimensional hydrofocusing of the sample fluid was analysed and demonstrated by cross sectional fluorescence imaging in good agreement with numerical simulations. Flow cytometry measurements have been performed by using 10μm particles. From the analysis of the fluorescence signals collected at each transit event we can confirm that the device was capable of creating a single-file particle stream. The results show that the device was capable of discriminating single microparticles with a good signal-to-noise ratio and a high throughput.

In this paper we demonstrate the ability to analyze a multiple of miniaturized bus-integrated sensor elements at high sensitivity from the superimposed complex overall spectrum by individual frequency modulation of optical microring resonators (MRR) fed by a single bus waveguide. The diverse sensor elements can be coated with biochemically selective adlayers (e.g., antibody molecules) to specifically promote the accumulation of target molecules on the MRR surface. Adhesion of target molecules results in an increase of the MRR resonance frequencies which can be measured at high sensitivity with a picometer accuracy. Readout of each MRR from the complex overall spectrum is performed by using phase sensitive lock-in detection to filter out the individual and selective response to external stimuli. We fabricated test arrays with 12 MRR elements based on silicon nitride material, each element integrated with a heater electrode for thermo-optical modulation of the MRR. A clear readout of the individual MRR by using a tuneable laser source is accomplished in a simple and reliable manner via lock-in detection despite strong overlap of the individual resonances. With our first results, we point out the large potential for multiplexed label-free detection of diverse biomolecular compounds.

We describe a photoluminescence-based, analyte-responsive sensor platform that is ~7 nm in diameter.

In the presented paper the optical system with converging spherical wave illumination for classification of bacteria species, is proposed. It allows for compression of the observation space, observation of Fresnel patterns, diffraction pattern scaling and low level of optical aberrations, which are not possessed by other optical configurations. Obtained experimental results have shown that colonies of specific bacteria species generate unique diffraction signatures. Analysis of Fresnel diffraction patterns of bacteria colonies can be fast and reliable method for classification and recognition of bacteria species. To determine the unique features of bacteria colonies diffraction patterns the image processing analysis was proposed. Classification can be performed by analyzing the spatial structure of diffraction patterns, which can be characterized by set of concentric rings. The characteristics of such rings depends on the bacteria species. In the paper, the influence of basic features and ring partitioning number on the bacteria classification, is analyzed. It is demonstrated that Fresnel patterns can be used for classification of following species: Salmonella enteritidis, Staplyococcus aureus, Proteus mirabilis and Citrobacter freundii. Image processing is performed by free ImageJ software, for which a special macro with human interaction, was written. LDA classification, CV method, ANOVA and PCA visualizations preceded by image data extraction were conducted using the free software R.

A new analytical method of high-selective detection of mycotoxins in food and feed are considered. A method is based on optical registration the changes of conduct of the electric polarized bacterial agents in solution at the action of the external gradient electric fields. Measuring are conducted in integrated electrode-optical cuvette of the special construction, which provides the photometric analysis of forward motion of the objects registration in liquid solution under act of the enclosed electric field and simultaneous registration of kinetics of change of electrical impedance parameters solution and electrode system.