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Results for “Microchip Analytical Procedures”

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At least 37 records · Page 2Linked to original sources

PubMedDate not supplied

Chemiluminescence micro-flow-injection analysis on a chip.

Chemiluminescence microflow-injection analysis (microFIA) systems on a chip have been developed. The technology of laser ablation was used to fabricate the microchannels on the polymethyl methacrylate (PMMA) chip. The three sampling structure, including double-tee sampling structure, microvalve sampling structure and injection pump with accurate time control, were used. The microcolumn for specific molecular recognition, including molecularly imprinted polymer, enzyme and bacteria, were used to enhance selectivity. These microFIA systems have been applied to clinical analysis, assessment of food safety, in vivo and real-time determination of drugs, and pharmacokinetics studies.

Animals↗

PubMedDate not supplied

Immunosorbent assay microchip system for analysis of human immunoglobulin G on MagnaBind carboxyl derivatized beads.

An immunosorbent assay system was integrated into a PMVS microchip. MagnaBind carboxyl derivatized beads were introduced into a microchannel, and then human immunoglobulin G (IgG) was bound to the bead surface in the microchannel of the chip. Immunoreaction was conducted in the microchannel for the bead-bounded antigen IgG with the antibody FITC-labelled IgG. On-chip detection was performed using a laser-induced fluorescence (LIF) system. The integration shortened the overall analysis time from 7 h to less than 40 min.

Antigens↗

PubMed2006

Recent advances of microfluidics in Mainland China.

The history and current status of research on microfluidics in China is summarized in this review. The recent representative contributions in this field by Chinese scientists are cited. A perspective on some trends in future development of this field in China is presented.

Animals↗

PubMed2006

Surface-enhanced laser desorption/ionization time-of-flight mass spectrometry serum protein profiling to identify nasopharyngeal carcinoma.

BACKGROUND: Diagnosis of nasopharyngeal carcinoma (NPC) at an early disease stage is important for successful treatment and improving the outcome of patients. The use of serum protein profiles and a classification tree algorithm were explored to distinguish NPC from noncancer. METHODS: Serum samples were applied to metal affinity protein chips to generate mass spectra by surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS). Protein peak identification and clustering were performed using the Biomarker Wizard software. Proteomic spectra of serum samples from 50 NPC patients and 54 noncancer controls were used as a training set and a classification tree with 6 distinct protein masses was generated by using Biomarker Pattern software. The validity of the classification tree was then challenged with a blind test set including another 20 NPC patients and 25 noncancer controls. RESULTS: The software identified an average of 93 mass peaks/spectrum and 6 of the identified peaks were used to construct the classification tree. The classification tree correctly determined 83% (123 of 149) of the test samples with 83% (58 of 70) of the NPC samples and 82% (65 of 79) of the noncancer samples. In a combination of the serum protein profiles with Epstein-Barr (EBV) nuclear antigen 1 (EBNA1 IgA) test, the diagnostic sensitivity and specificity were increased to 99% and 96%, respectively. CONCLUSIONS: The results suggest that SELDI-TOF-MS serum protein profiles could discriminate NPC from noncancer. The combination of serum protein profiles with an EBV antibody serology test could further improve the accuracy of NPC screening.

Adult↗

PubMed2006

Microtransponders, the miniature RFID electronic chips, as platforms for cell growth in cytotoxicity assays.

BACKGROUND: An electronic radio frequency (RF) microchip, the microtransponder (MTP), has been developed as a platform for assays in the fields of genomics and proteomics. Upon activation by light, each MTP provides a unique RF identification (ID) signal that matches a chip to the specific biological material attached to it. The MTP is powered by a photocell and has an antenna that transmits the signal. The aim of the present study was to explore utility of MTPs as a platform for cell growth in cytotoxicity assays. METHODS: The MCF-7, MCF-116, A549, or T-24 cells growing on MTPs placed in petri dishes or slide chambers were cultured untreated or exposed to antitumor drugs topotecan, mitoxantrone, or onconase for up to 4 days. Their attachment to- and growth on- MTPs was assessed by fluorescence microscopy and laser scanning cytometry (LSC) and compared with growth on the dish surface in the MTP neighborhood. The MTPs were fixed in ethanol, stained with propidium iodide (PI), and interrogated in flow in the instrument capable to rapidly (up to 103 MTPs/s) identify their ID signal and measure fluorescence. RESULTS: The cells plated on MTPs exhibited similar attachment properties to those plated in culture dishes. When measured by LSC, they had similar mitotic activity, growth rate, and cell cycle distributions as the cells adhering to the culture dish in the neighborhood of MTPs. The fluorescence intensity of MTPs provided information about the cell number per MTP, which made it possible to assess cell growth rate and monitor the cytostatic/cytotoxic effects of the tested drugs. CONCLUSIONS: The MTP-based system holds promise for the multiplexed cell assays in which numerous different cell lines can be screened for their growth rate or sensitivity while exposed to particular agents in the same vessel. Other advantages of the system are the rapidity of the screening and a very large number of ID codes. Because many cell lines/types can be assayed in a single dish, the system also offers cost savings on tissue culture reagents.

Adenocarcinoma↗

PubMed2004

Integrated on-chip derivatization and electrophoresis for the rapid analysis of biogenic amines.

We demonstrate the monolithic integration of a chemical reactor with a capillary electrophoresis device for the rapid and sensitive analysis of biogenic amines. Fluorescein isothiocyanate (FITC) is widely employed for the analysis of amino-group containing analytes. However, the slow reaction kinetics hinders the use of this dye for on-chip labeling applications. Other alternatives are available such as o-phthaldehyde (OPA), however, the inferior photophysical properties and the UV lambdamax present difficulties when using common excitation sources leading to a disparity in sensitivity. Consequently, we present for the first time the use of dichlorotriazine fluorescein (DTAF) as a superior in situ derivatizing agent for biogenic amines in microfluidic devices. The developed microdevice employs both hydrodynamic and electroosmotic flow, facilitating the creation of a polymeric microchip to perform both precolumn derivatization and electrophoretic analysis. The favorable photophysical properties of the DTAF and its fast reaction kinetics provide detection limits down to 1 nM and total analysis times (including on-chip mixing and reaction) of <60 s. The detection limits are two orders of magnitude lower than current limits obtained with both FITC and OPA. The optimized microdevice is also employed to probe biogenic amines in real samples.

Biogenic Amines↗

PubMed2004

Determination of SARS-coronavirus by a microfluidic chip system.

We have developed a new experimental system based on a microfluidic chip to determine severe acute respiratory syndrome coronavirus (SARS-CoV). The system includes a laser-induced fluorescence microfluidic chip analyzer, a glass microchip for both polymerase chain reaction (PCR) and capillary electrophoresis, a chip thermal cycler based on dual Peltier thermoelectric elements, a reverse transcription-polymerase chain reaction (RT-PCR) SARS diagnostic kit, and a DNA electrophoretic sizing kit. The system allows efficient cDNA amplification of SARS-CoV followed by electrophoretic sizing and detection on the same chip. To enhance the reliability of RT-PCR on SARS-CoV detection, duplex PCR was developed on the microchip. The assay was carried out on a home-made microfluidic chip system. The positive and the negative control were cDNA fragments of SARS-CoV and parainfluenza virus, respectively. The test results showed that 17 positive samples were obtained among 18 samples of nasopharyngeal swabs from clinically diagnosed SARS patients. However, 12 positive results from the same 18 samples were obtained by the conventional RT-PCR with agarose gel electrophoresis detection. The SARS virus species can be analyzed with high positive rate and rapidity on the microfluidic chip system.

Autopsy↗

PubMed2004

Separation of proteins on surface-modified poly(dimethylsiloxane) microfluidic devices.

Separation and detection of proteins have been realized on nonionic surfactant-modified poly(dimethylsiloxane) (PDMS) microfabricated devices with end-column amperometric detection. The hydrophobic PDMS channels are turned into hydrophilic ones after being modified with Brij35 and facilitate the separation of proteins. The coating can remarkably reduce the adsorption of large protein molecules and is stable in the range of pH 6-12. The detection of proteins in such channels needs less rinsing time and thus efficiency is raised. Even large molecules of proteins can also be detected with better reproducibility and enhanced plate numbers. The relative standard deviation (RSD) of the migration time for glucose oxidase (GOD) is 2.2% (n = 19). Separation of GOD and myoglobin has been developed in modified channels. Predominant operational variables, such as the coating conditions, the concentration of surfactant and buffer, are studied in detail.

Dimethylpolysiloxanes↗

PubMed2004

Genotyping by dynamic heating of monolayered beads on a microheated surface.

A miniaturized bead-based dynamic allele-specific hybridization (DASH) approach for single-nucleotide polymorphism analysis is presented. Chips with integrated heater and temperature sensors for open-surface DNA analysis were microfabricated. Microcontact printing using a poly(dimethylsiloxane) (PDMS) stamp was employed to create monolayers of immobilized beads on the surface of the chip. This chip allows fast, well-controllable temperature ramping. The temperature distribution was homogeneous over the entire heater area. All three possible variants of an SNP site of a synthesized oligonucleotide were accurately scored using the bead-based DASH approach. Our assay has a nonoptimized temperature ramping rate of 4 degrees C-6 degrees C/min compared to earlier reported values of 2 degrees C-3 degrees C/min, thereby reducing the total analysis time by a factor of 2. Reliable DASH measurement data from areas as small as 12 x 13 microm was achieved. Our bead-based DASH approach has enabled a dramatic volume reduction and is a step towards developing a cost-effective high-throughput DASH method on arrays of single beads.

Alleles↗

PubMed2004

New directions of miniaturization within the biomarker research area.

An overview of the current trends within protein expression profiling is given where multidimensional separation of both gel and liquid phase techniques linked to mass spectrometry is viewed as a major route in the global proteome mapping. A clear trend in these biochemical developments is the effort to sequence and identify low-abundant protein expressions where assay miniaturization and integrated sample processing play a central role. Two areas of miniaturization within the proteomics field are addressed: (i) sample cleanup and enrichment, and (ii) silicon microstructure developments for protein chip microarrays.

Animals↗

PubMed2004

Capillary electrophoresis of inorganic ions: an update.

This review as a sequel of three earlier similar reports gives a summary of the progress and significant methodological developments, starting from 2002, in the use of capillary electrophoresis (CE) for inorganic ion analysis. As substantiated by the illustrative number of relevant references, improvements in sensitivity achieved both in and outside a CE system, advances in manipulating the separation selectivity, novel hardware configurations, and system performance innovations are continually being reported over the review period. Specifically viewed are the recent advancements in elemental (bio)speciation analysis, which remains one of the most fertile areas of CE research, as well as in three recently booming research topics: contactless conductivity detection, separations on microchips, and transient isotachophoretic preconcentration. A state-of-the-art picture of technique's potentialities within the field of interest presented here demonstrates that CE has become recognized and is growing in acceptance as a reliable alternative to traditional analytical methods such as high-performance liquid chromatography (HPLC).

Electrophoresis, Capillary↗

PubMed2004

Recent advances in chiral separation principles in capillary electrophoresis and capillary electrochromatography.

This review summarizes recent developments in chiral separation in capillary zone electrophoresis (CZE), electrokinetic chromatography (EKC), and capillary electrochromatography (CEC) covering literature published since the year 2000. New chiral selectors and innovative approaches for CE and CEC are introduced. Recent progress in column technology for CEC is highlighted and the development of new chiral stationary phases is discussed. This review is not dedicated to list applications but will focus on new developments.

Chromatography, Liquid↗

PubMed2005

Microaffinity purification of proteins based on photolytic elution: toward an efficient microbead affinity chromatography on a chip.

A bead affinity chromatography system, which was based on the photolytic elution method, was integrated into a glass-silicon microchip to purify specific target proteins. CutiCore beads, which were coupled with a photo-cleavable ligand, such as biotin and an RNA aptamer, were introduced into a filter chamber in the microchip. The protein mixture containing target protein labeled with fluorescein isothiocyanate (FITC) was then passed through the packed affinity beads in the microchamber by pressure-driven flow. During the process, the adsorbed protein on the bead was monitored by fluorescence. The concentrated target protein on the affinity bead was released by simple irradiation with UV light at a wavelength of 360 nm, and subsequently eluted with the phosphate buffer flow. The eluted target protein was quantitatively detected via the fluorescence intensity measurements at the downstream of the capillary connected to the outlet of the microchip. The microaffinity purification allowed for a successful method for the identification of specific target proteins from a protein mixture. In addition, the feasibility of this system for use as a diagnosis chip was demonstrated.

Biotin↗

PubMed2005

A microfabricated capillary electrophoresis chip with multiple buried optical fibers and microfocusing lens for multiwavelength detection.

We present a new microfluidic device utilizing multiwavelength detection for high-throughput capillary electrophoresis (CE). In general, different fluorescent dyes are only excited by light sources with appropriate wavelengths. When excited by an appropriate light source, a fluorescent dye emits specific fluorescence signals of a longer wavelength. This study designs and fabricates plastic micro-CE chips capable of performing multiple-wavelength fluorescence detection by means of multimode optic fiber pairs embedded downstream of the separation channel. For detection purposes, the fluorescence signals are enhanced by positioning microfocusing lens structures at the outlets of the excitation fibers and the inlets of the detection fibers, respectively. The proposed device is capable of detecting multiple samples labeled with different kinds of fluorescent dyes in the same channel in a single run. The experimental results demonstrate that various proteins, including bovine serum albumin and beta-casein, can be successfully injected and detected by coupling two light sources of different wavelengths to the two excitation optic fibers. Furthermore, the proposed device also provides the ability to measure the speed of the samples traveling in the microchannel. The developed multiwavelength micro-CE chip could have significant potential for the analysis of DNA and protein samples.

Electrophoresis, Capillary↗

PubMed2005

Comparison of surfactants for dynamic surface modification of poly(dimethylsiloxane) microchips.

In the present report, the use of negatively charged surfactants as modifiers of the background electrolyte is reported using poly(dimethylsiloxane) (PDMS) microchips. In particular, the use of anionic surfactants, such as sodium dodecyl sulfate, phosphatidic acid, and deoxycholate, was studied. When surfactants were present in the run buffer, an increase in the electroosmotic flow (EOF) was observed. Two additional effects were also observed: (i) stabilization of the run-to-run EOF, (ii) an improvement in the electrochemical response for several biomolecules. In order to characterize the analysis conditions, the effects of different surfactant, electrolyte, and pH were studied. EOF measurements were performed using either the current monitoring method or by detection of a neutral molecule. The first adsorption/desorption kinetics studies are also reported for different surfactants onto PDMS. The separation of biologically important analytes (glucose, penicillin, phenol, and homovanillic acid) was improved decreasing the analysis time from 200 to 125 s. However, no significant changes in the number of theoretical plates were observed.

Adsorption↗

PubMed2006

Fluorescence imaging of sample zone narrowing and dispersion in a glass microchip: the effects of organic solvent (acetonitrile)-salt mixtures in the sample matrix and surfactant micelles in the running buffer.

A mismatch in the EOF velocities between the sample zone and running buffer region is known to generate pressure-driven, parabolic flow profile of the sample plug in electrokinetic separation systems. In the present study, video fluorescence microscopy was employed to capture real-time dynamics of the sample plug (containing fluorescein as the probe molecule) in a discontinuous conductivity system within a glass microchip, in which the sample matrix consisted of a mixture of ACN and salt (NaCl), and the running buffer contained sodium cholate (SC) micelles as the pseudo-stationary phase (i.e., performing "ACN stacking" in the mode of MEKC). Upon application of the separation voltage, the video images revealed that zone narrowing and broadening of the probe molecules occurred as the sample plug headed toward the cathode during the initial time period, probably resulting in part from the stacking/sweeping, and destacking of the SC micelles at the boundaries between the sample zone and running buffer. Interestingly, a second sample zone narrowing event can be observed as the sample plug moved further toward the cathode, which could be attributed to the sweeping of the slower moving probe molecules by the faster moving SC micelles that originated from the anode. This phenomenon was studied as a function of pH, sample plug length, as well as the concentration of organic solvent and salt in the sample matrix. The data suggested that the presence of large amounts of an organic solvent (such as ACN or methanol) and salts in the sample matrix not only induces sample dispersion due to the formation of a pressure-driven (hydrodynamic) flow, but may also lead to the formation of a double sample zone narrowing phenomenon by altering the local EOF dynamics within the separation system.

Acetonitriles↗

PubMed2004

Reduction of organic dyes in matrix-assisted laser desorption/ionization and desorption/ionization on porous silicon.

Reduction of analytes in matrix-assisted laser desorption/ionization (MALDI) often obscures the actual determination of molecular structure. To address the redox reactions in laser desorption/ionization processes, the organic dyes Methylene Blue, Janus Green B, Crystal Violet and Rhodamine B were analyzed by MALDI or by desorption/ionization on porous silicon (DIOS). Susceptibility to reduction in MALDI was dependent on both the reduction potentials of analytes and the molar ratio of analyte to matrix molecules. Addition of Cu(II) ions as an electron scavenger suppressed the reduction of Methylene Blue in MALDI. The results suggested that electron transfer to analytes from the sample target and/or from the matrix contributed to the reduction. In DIOS, the reductions of organic dyes were more prominent than in MALDI, and were not prevented by Cu(II) ion doping, probably due to direct contact of the analytes with silicon which had little electric resistance.

Azo Compounds↗

PubMed2005

Validating regulatory-compliant wide dynamic range bioanalytical assays using chip-based nanoelectrospray tandem mass spectrometry.

Automated chip-based infusion nanoelectrospray ionization coupled to tandem mass spectrometry (nanoESI-MS/MS) was used to validate a bioanalytical assay conforming to United States Food and Drug Administration (FDA) regulatory guidelines and Good Laboratory Practices (GLP). Reboxetine was used as the analyte fortified in dog plasma along with an analog internal standard (IS). The best nanoESI response for reboxetine was observed with 90% acetonitrile (ACN)/water without any mobile phase modifiers. The analyte and IS were extracted from dog plasma samples by liquid-liquid extraction (LLE). The supernatant was concentrated to dryness and redissolved in 90% ACN/water for nanoESI. Selected reaction monitoring (SRM) data were collected for all samples to generate ion current profiles with a base width of approximately 20 s. Selectivity experiments showed no interferences in blank plasma samples. Interferences as a result of in-source collision-induced dissociation of metabolites were not an issue due to the previously documented metabolism of reboxetine. Matrix suppression was evaluated across multiple lots of dog plasma as well as over different animal species (rabbit, rat, mouse) and different anticoagulants (heparin, EDTA). Matrix suppression ranged from approximately 30-60% across the different lots, species etc.; however, in all instances, the analyte and the IS were suppressed by similar amounts, suggesting the similarity in ionization properties between the two. A three-batch validation was performed (each batch consisting of four different concentrations, six replicates of each concentration) and demonstrated inter-assay accuracy (% relative error; RE) of less than +/-8% and an inter-assay precision (% relative standard deviation; RSD) of less than 7%, thus meeting regulatory guidelines. A comparison of analyses by nanoESI-MS/MS and liquid chromatography coupled to tandem mass spectrometry (LC/MS/MS) showed that nanoESI-MS/MS had a greater slope for the calibration standard curve compared to LC/MS/MS, indicating greater sensitivity for the former technique. It is also noteworthy that the amount of sample infused during nanoESI-MS/MS was approximately 80-fold less compared to the amount of sample injected during LC/MS/MS. The absence of carryover (attributed to the lack of a common fluid path) in the nanoESI technique enabled the extension of the assay linear dynamic range to 500,000-fold, and the possibility of analyzing samples in a single batch without the need for re-analysis of samples with high concentrations. This technology offers the possibility for increased throughput for studies supporting drug development by providing fast data turnaround for assays conforming to regulatory guidelines and GLPs.

Animals↗