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Characterizing permeability of the endothelium that lines blood vessels and heart valves provides fundamental physiological information and is required to evaluate uptake of drugs and other biomalecules. However, current techniques used to measure permeability, such as Transwell insert assays, do not account for the recognized effects of fluid flow-induced shear stress on endothelial permeability or are inherently low-throughput. Here we report a novel on-chip technique in a two-layer membrane-based microfluidic platform to measure real-time permeability of endothelial cell monolayers on porous membrsnes. Bovine serum albumin (a model protein) conjugated with fluoreseein isothiocyanate was delivered to an upper microchannel by pressure-driven flow and was forced to permeate a poly(ethylene terephthalate) membrane into a lower microchannel, where it was detected by laser-induced fluorescence. The concentration of the permeate at the point of detection varied with channel flow rates in agreement to less than 1% with theoretical analyses using a pore flow model. On the basis of the model, a sequential flow rate stepping scheme was developed and applied to obtain the permeability of cell-free and cell-bound membrane layers. This technique is a highly sensitive, novel microfluidic approach for measuring endothelial permeability in vitro, and the use of micrometer-sized channels offers the potential for parallelization and increased throughput compared to conventional shear-based permeability measurement methods. 10.1012/ac901560w

Buckmlnsterfullerene ([C.sub.60) as a primary ion for secondary ion mass spectrometry (SIMS) has shown many benefits over classical SIMS sources in the analysis of large organic molecules including many of biological significance. One constraint has been the limited focusing capabilities of the [C.sub.60] beam. Although this could be circumvented by using beam size limiting apertures at the cost of beam current, high-resolution imaging using conventional time-of-flight (TOF) instruments has been challenging and time-consuming. We present a method in which we combine the use of an unfocused [C.sub.60.sup.+] beam with an ion optical microscope. A delay line detector is used to obtain fully resolved hyperspectral data sets that contain both the full mass spectral and the localization information. The obtained image resolving power is 4[micro]m at a pixel size of 250 nm. Microscope mode [C.sub.60.sup.+] imaging was shown to resolve micrometer-scale features in a combined polymer-tissue sample. Our new approach demonstrates high-quality SIMS imaging using the full [C.sub.60.sup.+] beam current. This results in equal or better resolving power at reduced acquisition speed. 10.1021/ac902587g

No single biomarker can accurately predict disease. An ideal biodetection technology should be capable of the quantitative, reproducible, and sensitive detection of a limited suite of such molecules. To this end, we have developed a multiplex biomarker assay for protective antigen and lethal factor of the Bacillus anthracis lethal toxin using semiconductor quantum dots as the fluorescence reporters on our waveguide-based biosensor platform. The platform is extendable to a wide array of biomarkers, facilitating rapid, quantitative, sensitive, and multiplex detection, better than achievable by conventional immunoassay. Our assay allows for the sensitive (limit of detection 1 pM each), specific (minimal nonspecific binding), and rapid (15 min) detection of these biomarkers in complex biological samples (e.g., serum). To address the issue of reproducibility in measurement and to increase our sample throughput, we have incorporated multichannel waveguides capable of simultaneous multiplex detection of biomarkers in three samples in quadruplicate. In this paper, we present the design, fabrication, and development of multichannel waveguides for the simultaneous detection of lethal factor and protective antigen in serum. Evaluation of the multichannel waveguide shows an excellent concordance with single-channel data and effective, simultaneous, and reproducible measurement of lethal toxins in three samples. 10.1021/ac901497g

Because of the premature failure of in-service soft-body armor containing the ballistic fiber poly[(benzo-[1,2-d: 5,4-d']-benzoxazole-2,6-diyl)-l,4-phenylene] (PBO), the Office of Law Enforcement Standards (OLES) at the National Institute of Standards and Technology (NIST) initiated a research program to investigate the reasons for this failure and to develop testing methodologies and protocols to ensure that these types of failures do not reoccur. In a report that focused on the stability of the benzoxazole ring that is characteristic of PBO fibers, Holmes, G. A.; Rice, K.; Snyder, C. R. J. Mater. Sci. 2006, 41, 4105-4116, showed that the benzoxazole ring was susceptible to hydrolytic degradation under acid conditions. Because of the processing conditions for the fibers, it is suspected by many researchers that residual phosphoric acid may cause degradation of the benzoxazole ring resulting in a reduction of ballistic performance. Prior to this work, no definitive data have indicated the presence of phosphoric acid since the residual phosphorus is not easily extracted and the processed fibers are known to incorporate phosphorus containing processing aids. Methods to efficiently extract phosphorus from PBO are described in lids article. Further, characterization determined flint the majority of the extractable phosphorus in PBO was attributed to the octyldecyl phosphate processing aid with some phosphoric acid being detected. Analysis by matrix assisted laser desorption ionization of model PBO oligomers indicates that the nonextractable phosphorus is attached to the PBO polymer chain as a monoaryl phosphate ester. The response of model aryl phosphates to NaOH exposure indicates that monoaryl phosphate ester is stable to NaOH washes used in the manufacturing process to neutralize the phosphoric acid reaction medium and to extract residual phosphorus impurities. 10.1021/ac901602x

A capillary electrophoresis coupled with electrochemiluminescent detection system (CE-ECL) was developed for the detection of polymerase chain reaction (PCR) amplicons. The ECL luminophore, tris(1,10-phenanthroline) ruthenium(II) (Ru[(phen).sub.3.sup.2+]), was labeled to the PCR primers before amplification. Ru[(phen).sub.3.sup.2+] was then introduced to PCR amplicons by PCR amplification. Eventually, the PCR amplicons were separated and detected by the homemade CE-ECL system. The detection of a typical genetically modified organism (GMO), Roundup Ready Soy (RRS), was shown as an example to demonstrate the reliability of the proposed approach. Four pairs of primers were amplified by multiple PCR (MPCR) simultaneously, three of which were targeted on the specific sequence of exogenous genes of RRS, and another was targeted on the endogenous reference gene of soybean. Both the conditions for PCR amplification and CE-ECL separation and detection were investigated in detail. Results showed that, under the optimal conditions, the proposed method can accurately identifying RRS. The corresponding limit of detection (LOD) was below 0.01% with 35 PCR cycles. 10.1021/ac901510s

A wavelet-neural network signal processing method has demonstrated ~10-fold improvement over traditional signal processing methods for the detection limit of various nitrogen and phosphorus compounds from the output of a thermionic detector attached to a gas chromatograph. A blind test was conducted to validate the lower detection limit. All 14 of the compound spikes were detected when above the estimated threshold, including all 3 within a factor of 2 above the threshold. In addition, two of six spikes were detected at levels of half the concentration of the nominal threshold. Another two of the six would have been detected correctly if we had allowed human intervention to examine the processed data. One apparent false positive in five nulls was traced to a solvent impurity, whose presence was subsequently identified by analyzing a solvent aliquot evaporated to 1% residual volume, while the other four nulls were properly classified. We view this signal processing method as broadly applicable in analytical chemistry, and we advocate that advanced signal processing methods should be applied as directly as possible to the raw detector output so that less discriminating preprocessing and postprocessing does not throw away valuable signal.

The autonomous pathogen detection system (APDS) is an automated, podium-sized instrument that continuously monitors the air for biological threat agents (bacteria, viruses, and toxins). The system has been developed to warn of a biological attack in critical or high-traffic facilities and at special events. The APDS performs continuous aerosol collection, sample preparation, and detection using multiplexed immunoassay followed by confirmatory PCR using real-time TaqMan assays. We have integrated completely reusable flow-through devices that perform DNA extraction and PCR amplification. The fully integrated system was challenged with aerosolized Bacillus anthracis, Yersinia pestis, Bacillus globigii, and botulinum toxoid. By coupling highly selective antibody-and DNA-based assays, the probability of an ADDS reporting a false positive is extremely low.

Dual Pt disk microelectrodes consisting of a 10-[micro]m distance sensor and a 50-[micro]m nitric oxide sensor were prepared. The 50-[micro]m electrode was modified with Ni(4-N-tetramethyl) pyridyl porphyrin enclosed in the polymer network of a negatively charged electrodeposition paint. This paint prevented the dissolution of the otherwise soluble porphyrin in the aqueous test medium due to charge interactions. It also denied negatively charged ions in the analyte solution access to the electrode surface by electrostatic repulsion, thereby preventing interference from anions such as nitrite, nitrate, and ascorbate. With the aid of a scanning electrochemical microscope, it was possible to use the distance sensor by recording the negative feedback effect on the reduction of molecular oxygen to 'guide' the nitric oxide sensor to various known distances from a layer of adherently growing human umbilical vein endothelial cells for the detection of nitric oxide released from the cells upon stimulation with bradykinin. The use of the distance sensor made it possible to preserve the integrity of the adherently growing cells concomitantly with the modified electrode by preventing the deterioration of the modifying layer during the distance adjustment step.

Leukocyte isolation from whole blood to study inflammation requires the removal of contaminating erythrocytes. Leukocytes, however, are sensitive to prolonged exposure to hyper/hypoosmotic solutions, temperature changes, mechanical manipulation, and gradient centrifugation. Even though care is taken to minimize leukocyte activation and cell loss during erythrocyte lysis, it is often not possible to completely avoid it. Most procedures for removal of contaminating erythrocytes from leukocyte preparations are designed for bulk processing of blood, where the sample is manipulated for longer periods of time than necessary at the single-cell level. Ammonium chloride-mediated lysis is the most commonly used method to obtain enriched leukocyte populations but has been shown to cause some activation and selective loss of certain cell types. The leukocyte yield and subsequent activation status of residual leukocytes after N[H.sub.4]Cl-mediated lysis have been shown to depend on the time of exposure to the lysis buffer. We have developed a microfluidic lysis device that deals with erythrocyte removal at nearly the single-cell level. We can achieve complete lysis of erythrocytes and ~ 100% recovery of leukocytes where the cells are exposed to an isotonic lysis buffer for less than 40 s, after which the leukocytes are immediately returned to physiological conditions. Theoretically, this process can be made massively parallel to process several milliliterss of whole blood to obtain a pure leukocyte population in less than 15 min.

Halogen-containing polymeric flame retardants in engineering thermoplastics have been analyzed by pyrolysis gas chromatography (Py-GC) with an atomic emission detector (AED) and a mass selective detector (MSD). The atomic emission detector is very effective in monitoring halogen-containing pyrolysates after Py-GC. The type of polymeric flame retardant used in thermoplastics can be identified by peak pattern recognition through an AED halogen element trace. In this study, a set of thermoplastic resins containing different brominated polymeric flame retardants have been studied. After generation of the pyrogram by Py-GC/AED, those pyrolysates produced from polymeric flame retardants were identified by MSD analysis of a complex total ion chromatogram (TIC) generated by the thermoplastic resins. This study demonstrated the effectiveness of the AED in the tasks of specific element monitoring and component pattern recognition. The advantages and disadvantages of using the AED are also discussed.

Chromatographers generally try to achieve good resolution of components in a mixture as a prerequisite to quantitation. Occasionally, multiple detectors are used to resolve and quantitate chromatographically overlapping components. In most cases, however, the detectors exhibit very high selectivity for the overlapping components; l.e., one detector is much more sensitive to a particular molecular structural feature contained in one of the species being analyzed. In some cases, one detector exhibits little or no response for a particular component in a mixture, and so multiple detectors often function in a complementary fashion. This paper describes an analytical method for quantitating chromatographically overlapping components using multiple detectors that exhibit comparable, although not identical, selectivity for the coeluting species. Equations are derived for quantitating two overlapping components using two detectors with either an internal or an external standard analytical method. Quantitation is possible even when the components exhibit complete overlap. The internal standard method is applied to the analysis of acrolein/proplonaldehyde mixtures using a tandem photoionization/flame ionization detector. Component concentrations are determined with a relative accuracy of 3.4% when both overlapping components are present at equal concentrations. When the components are present at different concentrations, greater relative accuracy is obtained for the more highly concentrated component. Regression techniques may be used to generalize the method to the analysis of N overlapping components using at least N different detectors., An analytical method for the quantification of chromatographically overlapping components by multiple detectors was developed. The resolution of propionaldehyde and acrolein by tandem photoionization/flame photoionization detection tested the applicability of the technique. Results indicate that relative accuracy of the determined component concentrations affected the presence of overlapping components in the concentrations.

A 2.45-GHz microwave-induced plasma gas chromatographic detector integrated with water-cooled capillary plasma and tangential flow torches was subjected to sensitivity and reflected power measurements in order to determine the plasma torch effect on the system's analytical capability. The limit-of-detection and sensitivity results favored a water-cooled capillary system design over tangential flow. In addition, the reflected-power measurement gave a detection limit of 0.6 ng C/s, while optical-emission detection gave the best limit-value of 0.3 ng C/s.