[Show abstract][Hide abstract] ABSTRACT: Hydrophobic sol-gel derived methylsilsesquioxane (MSQ) was compared to wax and alkylketene dimer (AKD) as barrier materials defining channels in paper-based microfluidic devices. While all three of the barrier types performed well with water, only the MSQ barriers were not breached by aggressive cell lysing solutions and surfactant solutions (SDS, CTAB, Triton X-100). The MSQ barriers also withstood glycerol, toluene and DMSO whereas all three barrier types were breached by alcohols. MSQ based devices could be prepared either by direct inkjet printing of the barriers or by base etching of MSQ impregnated paper. The functionality of MSQ based devices was demonstrated by formatting a colormetric sensor for Escherichia coli.
Lab on a Chip 12/2013; 14(4). DOI:10.1039/c3lc51313k · 6.12 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A thin-film octadecyl (C18)-silica glass coating was developed as the extraction phase for an automated 96-blade solid-phase microextraction (SPME) system coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Various factors (e.g., sol-gel composition and aging time, coating preparation speed, coating thickness, and drying conditions) affecting the quality of C18-silica glass thin-film coating were studied and optimized. The results showed that the stability and durability of the coating are functions of the coating thickness and drying conditions. Coating thickness is controlled by sol-gel composition, aging time and the withdrawal speed in the dipping method. Automated sample preparation was achieved using a robotic autosampler that enabled simultaneous preparation of 96 samples in a 96-well plate format. Under the optimum SPME conditions the proposed system requires a total of 140 min for preparation of all 96 samples (i.e., 30 min preconditioning, 40 min equilibrium extraction, 40 min desorption and 30 min carry over step). The performance of the C18-silica glass 96-blade SPME system was evaluated for high-throughput analysis of benzodiazepines from phosphate-buffered saline solution (PBS) and human plasma, and the reusability, repeatability, and validity of the system were evaluated. When analysing spiked PBS and human plasma, the inter-blade reproducibility for four benzodiazepines was obtained in the ranges of 4-8% and 9-11% RSD (relative standard deviation), respectively, and intra-blade reproducibility were in the ranges of 3-9% and 8-13% RSD, respectively. The limits of detection and quantitation for plasma analysis were in the ranges of 0.4-0.7 ng/mL and 1.5-2.5 ng/mL for all four analytes.
Journal of Chromatography A 11/2011; 1246:2-8. DOI:10.1016/j.chroma.2011.11.030 · 4.17 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The last two decades have seen a revolution in the area of sol–gel-derived materials as media for the immobilization of biomolecules for biosensor fabrication. Such materials are suitable for the entrapment of a range of biomolecules, from enzymes to antibodies and even functional nucleic acids (FNA) such as aptamers and DNA enzymes. Recent progress in the development of “protein friendly” sol–gel processing methods has allowed these materials to be utilized as components of numerous biosensors, using delicate biomolecules such as luciferease and kinases, or even membrane-bound receptors as biorecognition elements. In addition, such materials have proven to be particularly versatile in the fabrication of biosensors, being amenable to methods such as dipcasting, contact printing, or even noncontact inkjet printing to form a bioselective coating on a range of substrates. In this review, we provide an overview of advances in biofriendly sol–gel processing methods developed in our research group and others, and we highlight accomplishments in the immobilization of both proteins and FNA within silica based materials. We then describe methods for interfacing biomolecule-doped materials to optical biosensors, with emphasis on fiber optic sensors, microarray-based multianalyte sensors and bioactive paper-based test strips. In each case, the material processing requirements for fabrication of different devices is emphasized. Finally, a brief perspective on potential future areas of research in the field of sol–gel based biocomposites is provided.
Chemistry of Materials 11/2011; 24(5):796–811. DOI:10.1021/cm202798e · 8.35 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Protein microarrays based on pin-printing of sol–gel-entrapped biomolecules have emerged as a potential tool to accelerate drug screening and discovery. However, while materials have recently been identified that are suitable for printing of high-density sol–gel-based microarrays, the ability to print arrays of delicate proteins such as kinases, and to assay their activity and inhibition on-array, has yet to be demonstrated. In this study, we have performed a criteria-based directed screen of sol–gel-based materials to identify compositions that are suitable for the fabrication of high-density, multikinase microarrays. Printable formulations were assessed for their compatibility with a fluorescent, phosphospecific dye used as an end-point indicator for on-array kinase assays, including an assessment of the effects of spot size (100 μm vs 400 μm) and slide surface chemistry on signal reproducibility. The combinations of materials, surfaces, and spot sizes that were found to be compatible with reproducible signal generation were evaluated for their ability to retain the activity of a range of kinases, which were co-entrapped with their respective substrates into the optimal sol–gel materials to produce microarrays. Ultimately, two material/surface combinations, from potentially thousands, were identified, one of which was used to produce a robust, highly active kinase microarray that could be used for qualitative screening as well as quantitative inhibition assays.
Chemistry of Materials 08/2011; 23(16):3685-3691. DOI:10.1021/cm2012389 · 8.35 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A fluorimetric acetylcholinesterase (AChE) assay was developed and characterized both in solution and with the enzyme entrapped in sol-gel-derived silica. The assay is based on a disulfide-thiol interchange reaction between the intramolecularly quenched dimeric dye BODIPY FL l-cystine and thiocholine generated by the AChE-catalyzed hydrolysis of acetylthiocholine (ATCh), which results in a brightly fluorescent monomeric product owing to the cleavage of the disulfide-coupled form of the dye. The new assay was validated by comparison with the Ellman assay performed under parallel conditions and was used in both kinetic and end point assays. The assay was extended to the fabrication of functional AChE microarrays using contact pin-printing of sol-gel-derived silica. A total of 392 sol-gel formulations were screened for gelation times and 192 of these were further evaluated for array fabrication on four different surfaces using a factor analysis approach. Of these, 66 sol-gel/surface combinations produced robust microarrays, while 26 sol-gel/surface combinations were identified that could produce highly active AChE microarrays. The Z' factor for the on-array assay using an optimal sol-gel/surface combination, which considers both signal variability and difference in signals between positive and negative controls, was determined to be 0.60, which is above the minimum level required for applicability to screening. By overprinting nanoliter volumes of solutions containing the dye, ATCh, and potential inhibitors, these microarrays could be used to screen two libraries of small molecules, one composed of newly synthesized alkaloids and another consisting of ∼1000 known bioactive compounds, both as discrete compounds and mixtures thereof, for activity against AChE. IC(50) values were obtained on microarrays for compounds showing significant inhibitory activity, demonstrating the utility of arrays for quantitative inhibition assays.
[Show abstract][Hide abstract] ABSTRACT: We present a novel way to prepare SPME fibers using a silicate entrapment of porous particles, followed by derivatization using classical organosilane chemistry. The fibers provide a good platform for on-fiber derivatization of desired extraction phases while providing porosity necessary for high extractions capacities. The porous network was created using potassium silicate and porous silica particles. Fibers derivatized using n-butyl, n-octyl, n-octadecyl and n-triacontyl groups were shown to extract benzodiazepines successfully. The coatings were determined to have an average thickness of ca. 8 microm, as determined by a scanning electron microscope, permitting equilibrium times as fast as 2 min. The fibers also showed very good ruggedness towards a vast range of solvents and prolonged use. It was determined that greater extraction efficiencies could be obtained using triacontyl as an extraction phase. The C18 and C30 fibers were also found to provide good linearity (>0.99) for the model analytes over two orders of magnitude, with limits of detection in the sub ng mL(-1) levels. C30 fibers were used to establish a correlation between structurally diverse beta-blockers and their literature reported Log P values. The C30 fibers provided a good correlation (R(2)=0.9255) between beta-blockers ranging in hydrophobicity from Log P(literature) 0.16-4.15 and their respective experimentally determined Log K(spme) values.
[Show abstract][Hide abstract] ABSTRACT: An easy-to-implement capillary coating strategy based on polymer entrapment in the network of polymerized silicate is described. In this manner, cationic polymers are tightly fixed onto the inner wall of the capillary for electroosmotic flow control without necessitating complex surface modification chemistries. The resulting coated capillary exhibited good stability over a wide range of pH, good reproducibility, strong endurance in more than 300 electrophoretic runs, and tolerance of commonly employed organic solvent additives in CE. Applications in CE-MS analysis of biologically important anions as well as sample enrichment are shown. Additionally, it was used as a durable base for attachment of multiple layers of charged polymers on the wall, via electrostatic interaction with the preceding layer. Thus, two novel types of highly stable coated capillaries, one with anodic EOF and the other cathodic, were developed.
[Show abstract][Hide abstract] ABSTRACT: Capillary electrophoresis (CE)-mass spectrometry (MS), as an analytical platform, has made significant contributions in advancing metabolomics research, if still limited up to this time. This review, covering reports published between 1998 and 2006, describes how CE-MS has been used thus far in this field, with the majority of the works dealing with targeted metabolite analyses and only a small fraction using it in the comprehensive context. It also discusses how some of the key features of CE-MS were exploited in selected metabolomic applications.
Journal of Chromatography A 11/2007; 1168(1-2):237-46; discussion 236. DOI:10.1016/j.chroma.2007.02.065 · 4.17 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We report an on-line sample preconcentration technique based on dynamic pH junction in capillary electrophoresis-mass spectrometry (CE-MS). For peptide analysis, the samples were dissolved in a solution with higher pH than the background solution (BGS), and were injected into the capillary as a long plug. The pH difference between the sample matrix and BGS caused changes in analytes' mobilities during electrophoresis, resulting in narrowing of their bands at the boundary. Around 550-1000-fold sensitivity enhancement could be achieved in terms of peak intensity without degrading peak shape and resolution. This technique is easy to perform and will be useful for peptide mass fingerprinting in protein analysis.
Journal of Chromatography A 06/2007; 1148(2):250-5. DOI:10.1016/j.chroma.2007.03.014 · 4.17 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Compared to chromatography-based techniques, the concentration limits of detection (CLOD) associated with capillary electrophoresis are worse, and these have largely precluded their use in many practical applications. To overcome this limitation, researchers from various disciplines have exerted tremendous efforts toward developing strategies for increasing the concentration sensitivities of capillary electrophoresis (CE) systems, via the so-called sample enrichment techniques. This review highlights selected developments and advances in this area as applied to the analyses of proteins and peptides in the last 5 years.
Journal of Chromatography B 10/2006; 841(1-2):88-95. DOI:10.1016/j.jchromb.2006.04.017 · 2.73 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A method based on the presence of a dynamic pH junction within the capillary to induce band narrowing for enhanced detection sensitivity for some peptides is presented. This technique is predicated on a sharp reduction in an analyte's migration velocity following a reversal of its electrophoretic direction from the acidic sample zone to the basic BGS zone. Larger-than-usual injection volumes of samples in relatively high-conductivity matrices were enabled, without degrading peak shape, resolution and efficiency. The size of the original sample plug was reduced by as much as 38-fold, and improvement in detector response in terms of peak height by as much as 124-fold was obtained. The effects of pH and concentration of the sample matrix, and the length of sample injection on the efficiency of the technique are discussed.
Journal of Chromatography A 07/2005; 1079(1-2):266-73. DOI:10.1016/j.chroma.2005.03.069 · 4.17 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Many researchers have invested considerable efforts toward improving capillary electrophoresis (CE)-mass spectrometry (MS) systems so they can be applied better to standard analyses. This review highlights the developments in CE-MS of proteins and peptides over the last five years. It includes the developments in interfaces, sample-enrichment techniques, microfabricated devices, and some applications, largely in capillary zone electrophoresis (CZE), capillary isoelectric focusing (CIEF) and capillary isotachophoresis formats.
[Show abstract][Hide abstract] ABSTRACT: Field-enhanced sample injection (FESI) was used to improve the concentration sensitivity of a capillary electrophoresis (CE)-mass spectrometry (MS) system with sheath flow configuration. Using some bioactive peptides, more than 3000-fold improvement in signal was obtained, permitting analysis in the low nM (fmol/microl) levels. The system was further evaluated for analysis of complex peptide mixtures by using low concentration tryptic digests of standard proteins. Rapid identification of the original protein was obtained by database searching using the observed molecular masses of the peptides, and by comparison of actual MS-MS spectra of selected peptides with the predicted fragmentation patterns.
Journal of Chromatography A 05/2004; 1032(1-2):203-11. DOI:10.1016/j.chroma.2003.10.038 · 4.17 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: On-line preconcentration by sweeping in micellar electrokinetic chromatography using mixed micelles of sodium dodecyl sulfate (SDS)-SB-12 is presented. Because of their large micelle radius, they permit increased partitioning of hydrophobic analytes into the core. In addition, they also possess lower negative surface charge relative to pure SDS micelles so anionic analytes can be retained better due to decreased electrostatic repulsion. As the efficiency of sweeping is predicated on the magnitude of retention factors, these advantages translated to better focusing. As much as a 370-fold improvement in detector response, in terms of peak height, was obtained for some neutral steroids, while about a 360-fold improvement was obtained for some phenol derivatives, which were previously not amenable to sweeping by pure SDS micelles.
Journal of Chromatography A 02/2003; 985(1-2):435-45. DOI:10.1016/S0021-9673(02)01654-0 · 4.17 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The separation and on-line preconcentration by sweeping of charged analytes in electrokinetic chromatography using a neutral pseudostationary phase is described. Under neutral or basic conditions, the electrophoretic migration of anionic analytes towards the anode is overcome by a high, cathode-directed electroosmotic flow; hence, they experience net migration towards the cathode, and the system is run at positive polarity mode. The separation and the retention factor, k, are dependent on both the analyte's electrophoretic mobility and its interaction with the pseudostationary phase. The versatility of the sweeping mechanism is then shown in this system. The charged analyte, prepared in a matrix free of the pseudostationary phase, penetrates the pseudostationary phase zone upon application of voltage. Analyte molecules are consequently accumulated and concentrated. As a demonstration, the separation and preconcentration of phenol derivatives using nonionic surfactants of the alkyl polyoxyethylene ether type (Brij 35 and Brij 58) yielded peak height enhancements up to 100-fold. The efficiency of sample stacking was also found to be improved with the use of a high viscosity background solution.
Journal of Chromatography A 01/2002; 939(1-2):99-108. DOI:10.1016/S0021-9673(01)01346-2 · 4.17 Impact Factor