| 1. |
- Gascón, Jordi, et al.
(författare)
-
Performance of two immunoassays for the determination of atrazine in sea water samples as compared with on-line solid phase extraction-liquid chromatography-diode array detection
- 1996
-
Ingår i: Analytica Chimica Acta. - : Elsevier BV. - 0003-2670. ; 330:1, s. 41-51
-
Tidskriftsartikel (refereegranskat)abstract
- Two immunoassay formats, magnetic particles-based assay (Atrazine RaPID assay and Atrazine High-Sensitivity RaPID assay) and microtiter plate based assay (Department of Entomology and Environmental Toxicology, University of California in Davis) were evaluated for the determination of atrazine in sea water samples. The results obtained were compared and validated with those obtained by using on-line solid phase extraction followed by liquid chromatography-diode array detection (on-line SPE-LC-DAD). The correlation between both techniques was good when analyzing levels of atrazine ranging from 0.01 to 5 μg/l in samples showing salt concentration values varying from 0 to 35 g/l and pH values from 2 to 10. One of these immunoassays (Atrazine High-Sensitivity RaPID assay) was employed to directly analyze atrazine in real estuarine and coastal water samples. The same samples were analyzed after filtration and C18 Empore disks extraction.
|
|
| 2. |
- Lutz, Mareike, 1967-, et al.
(författare)
-
Effects of different additives on a tyrosinase based carbon paste electrode
- 1995
-
Ingår i: Analytica Chimica Acta. - Amsterdam : Elsevier. - 0003-2670 .- 1873-4324. ; 305:1-3, s. 8-17
-
Tidskriftsartikel (refereegranskat)abstract
- The influence of a number of solid and chemical additives on the sensitivity and operational stability of a tyrosinase carbon paste electrode was studied. Cyclic voltammograms were run of the electrochemically active catechol/o-quinone couple on unmodified and additive modified carbon paste electrodes without tyrosinase. This was done in order to study the influence of these additives on the pure electrochemistry of the carbon paste. The influence on the total system (additive and enzyme modified carbon paste electrode) was studied in the flow injection mode. In some instances a dramatic improvement of the direct electron transfer of the catechol/o-quinone couple was obtained with both solid and chemical additives included in the carbon paste. A similar improvement of biosensor sensitivity in the flow injection mode was obtained with most chemical additives whereas the solid additives had a negative impact on biosensor sensitivity. The results obtained in this work indicate that these additives influence the purely electrochemical processes at the carbon paste and/or the performance of the enzyme in the carbon paste environment. How and why these additives can possibly influence the biosensor performance are discussed. © 1995.
|
|
| 3. |
- Marko-Varga, György, et al.
(författare)
-
Effect of HY-Zeolites on the Performance of Tyrosinase-Modified Carbon Paste Electrodes
- 1996
-
Ingår i: Electroanalysis. - Weinheim : Wiley-VCH Verlagsgesellschaft. - 1040-0397 .- 1521-4109. ; 8:12, s. 1121-1126
-
Tidskriftsartikel (refereegranskat)abstract
- The dependence of electrode response on additive properties in enzyme-modified carbon paste was studied. Four different HY-zeolite powders, dealuminated to different extents and characterized by both Si/Al ratio and hydrophilicity, were used as the carbon paste modifiers. The enzyme tyrosinase used in biosensors for the detection of catechol and other phenolic compounds was chosen as the model system for the construction of a composite carbon paste biosensor incorporating different HY-zeolites as additives. Tyrosinase was trapped on the HY-zeolite particles from a buffer solution, dried and mixed with graphite powder and a pasting oil. It was found that by incorporating HY-zeolites into the carbon paste the heterogeneous reaction rate of catechol redox conversion and the signal response for catechol were increased. In the latter case a higher response was observed for increased hydrophilicity, i.e., decreased Si/Al ratio of the HY-zeolite. The carbon paste/solution interface is considered to be an aqueous/organic phase and the characteristics of the enzyme-modified carbon paste electrode are related to theories, explaining enzymatic catalysis in organic solvents.
|
|
| 4. |
- Torto, N, et al.
(författare)
-
Monitoring of enzymatic hydrolysis of starch by microdialysis sampling coupled on-line to anion exchange chromatography and integrated pulsed electrochemical detection using post-column switching
- 1997
-
Ingår i: Biotechnology and Bioengineering. - 1097-0290. ; 56:5, s. 546-554
-
Tidskriftsartikel (refereegranskat)abstract
- A quantitative evaluation of the hydrolysis of wheat starch using Termamyl, a thermostable alpha-amylase (endo-l,4-alpha-D-glucan, glucanohydrolase; EC 3.2.1.78), is reported. Data from the monitoring of the hydrolysis of wheat starch indicated that, after 1 h, glucose and maltooligosaccharides up to DP 7 were the main hydrolysis products and thus enabled optimization of a liquefication step during the production of L-lactic acid. The monitoring system used, both in the on- and off-line mode, was based on continuous flow microdialysis sampling (CFMS) coupled to anion exchange chromatography and integrated pulsed electrochemical detection (IPED). A microdialysis probe equipped with a 5-mm polysulfone (SPS 4005) membrane, with a molecular-weight cut-off of 5 kDa, was used to sample the hydrolysis products of native wheat starch at 90 degrees C. Characteristic fingerprint separations were achieved by anion exchange chromatography after enzymatic hydrolysis. Post-column switching improved the detection and, consequently, also quantification of the hydrolysates as fouling of the electrode could be reduced. Maltooligosaccharide standards were used for quantification and to verify the elution of the hydrolysates by spiking the off-line samples. (C) 1997 John Wiley & Sons, Inc.
|
|
| 5. |
- Torto, Nelson, et al.
(författare)
-
On-line quantitation of enzymatic mannan hydrolysates in small-volume bioreactors by microdialysis sampling and column liquid chromatography-integrated pulsed electrochemical detection
- 1996
-
Ingår i: Journal of Chromatography A. - : Elsevier BV. - 0021-9673. ; 725:1, s. 165-175
-
Tidskriftsartikel (refereegranskat)abstract
- Quantitative aspects of on-line microdialysis sampling are investigated with respect to sampling in small-volume bioreactors. Three modes of microdialysis sampling; continuous-flow microdialysis sampling (CFMS), stopped-flow microdialysis sampling with continuous stirring, and stopped-flow microdialysis sampling with stopped stirring were investigated as a possible means for studying bioprocesses. The hydrolytic properties of a well characterised endomannanase from Aspergillus niger were studied using 0.01% ivory nut mannan as substrate in a 5-ml reaction vessel. On-line sampling was achieved using a microdialysis probe fitted with an SPS 6005 membrane. Stopped-flow microdialysis sampling was found to give the least analyte depletion and thus used for quantitation of the enzymatic hydrolysates. However, CFMS can be used when analyte depletion is not significant (large-volume reactor). Hydrolysis of ivory nut mannan gave mainly mannobiose and mannotriose in almost equal amounts, which is consistent with an endo-wise hydrolysis. The concentrations of mannose and mannopentaose did not change significantly over the monitoring period, however, that of mannotetraose increased gradually up to 11 h where it starts to decrease.
|
|
| 6. |
- Önnerfjord, Patrik, et al.
(författare)
-
A flow immunoassay for studies of human exposure and toxicity in biological samples
- 1998
-
Ingår i: Journal of Molecular Recognition. - 0952-3499. ; 11:1-6, s. 182-184
-
Tidskriftsartikel (refereegranskat)abstract
- This paper describes a heterogeneous competitive flow immunoassay with a high sample throughput which can be used for the screening of smaller analytes in various samples. The method is based on off-line incubation of the analyte (Ag), a fluorescent labelled tracer (Ag*) and the corresponding antibody (Ab). The separation of bound (Ab-Ag*) and free tracer (Ag*) is based on a size exclusion and reversed phase mechanism utilizing a restricted access (RA) column. The column traps the free unbound tracer (Ag*) in its hydrophobic (C18) inner cavity but excludes the large Ab-Ag* complex, which is passed on and measured by the fluorescence detector. The flow immunoassay was developed using the triazine herbicide atrazine as a model compound owing to its human toxicity and widespread use. A sample throughput of 80 samples per hour and a detection limit of 300 pg ml-1 in water were obtained. Urine samples were successfully applied for direct injections into the flow system, while for human plasma samples an additional clean-up step using solid phase extraction was efficiently included where pure extract is obtained with the highly stable and biocompatible extracting column material. The resulting detection limits for atrazine in plasma and water samples using this clean-up and trace enrichment procedure were found to be 2 ng ml-1 and 20 pg ml-1 respectively.
|
|
| 7. |
- Önnerfjord, Patrik, et al.
(författare)
-
High sample throughput flow immunoassay utilising restricted access columns for the separation of bound and free label
- 1998
-
Ingår i: Journal of Chromatography A. - 0021-9673. ; 800:2, s. 219-230
-
Tidskriftsartikel (refereegranskat)abstract
- A flow immunodetection system with high sample throughput capacity is described for the screening of various analytes. The immunochemical detection principle is based on the chromatographic separation of the formed immunocomplex (AbAg or AbAg*) and the free antigen (Ag) by a restricted access (RA) column, utilising size-exclusion and reversed-phase mechanism. A fluorescein labelled analyte (Ag*) was used in the competitive assay format with fluorescence detection. The speed and simplicity of the assay were the greatest advantages, allowing measurement of the analyte to be carried out in less than 1 min. The biocompatibility and capacity of the restricted access material allowed multiple injections of up to 5000, without any breakthrough of the fluorescent tracer molecule and thus need for regeneration. The flow immunoassay was developed using the well-known atrazine herbicide and some transformation products as model compounds, due to their human toxicity and widespread use. The sample throughput was 80 samples per hour and the detection limits were 1.4 nM (300 pg/ml) for atrazine (Ab I) and 2.3 nM (500 pg/ml) for the sum of triazines (Ab II-III). Different sample matrices, PBS buffer, creek water, and urine were successfully applied in the flow system without the need for any sample handling step. For plasma samples an additional clean-up step using solid-phase extraction had to be included. The resulting detection limits for atrazine in plasma and water samples using this clean-up and trace enrichment procedure were found to be 2 ng/ml and 20 pg/ml, respectively. The analysis could be performed at a sample throughput rate of 400 per 6-h working shift.
|
|
| 8. |
- Önnerfjord, Patrik, et al.
(författare)
-
Homogeneous sample preparation for automated high throughput analysis with matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry.
- 1999
-
Ingår i: Rapid Communications in Mass Spectrometry. - 0951-4198 .- 1097-0231. ; 13:5, s. 315-22
-
Tidskriftsartikel (refereegranskat)abstract
- This work presents a simple method for obtaining homogeneous sample surfaces in matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOFMS) for the automated analysis of peptides and proteins. The sample preparation method is based on applying the sample/matrix mixture onto a pre-deposited highly diluted matrix spot. The pre-deposited crystals act as seeds for the new sample containing crystals which become much smaller in size and more evenly distributed than with conventional methods. This 'seed-layer' method was developed, optimised and compared with the dried-droplet method using peptides and proteins in the 1000-20,000 Da range. The seed-layer method increases the surface homogeneity, spot to spot reproducibility and sample washability as compared with the commonly used dried-droplet method. This methodology is applicable to alpha-cyanohydroxycinnamic acid, sinapinic acid and ferulic acid, which all form homogeneous crystal surfaces. Within-spot variation and between-spot variation was investigated using statistics at a 95% confidence level (n = 36). The statistical values were generated from more than 5000 data points collected from 500 spectra. More than 90% of the sample locations results in high intensity spectra with relatively low standard deviations (RSDs). Typically obtained data showed an RSD of 19-35% within a sample spot as well as in-between spots for proteins, and an RSD of < or = 50% for peptides. Linear calibration curves were obtained within one order of magnitude using internal calibration with a point-RSD of 3% (n = 10). The sample homogeneity allows mass spectra (average of 16 laser shots) to be obtained on each individual sample within 15 sec, whereby a 100 spot target plate can be run in 25 min. High density target plates using the seed-layer method were prepared by spotting approximately 100 picoliter droplets onto the target, resulting in sample spots < or = 500 microns in diameter using a flow-through piezo-electric micro-dispenser. By using this automated sample preparation step lower standard deviations are obtained in comparison to manually prepared samples.
|
|
| 9. |
- Önnerfjord, Patrik, et al.
(författare)
-
Picoliter sample preparation in MALDI-TOF MS using a micromachined silicon flow-through dispenser
- 1998
-
Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 0003-2700 .- 1520-6882. ; 70:22, s. 4755-4760
-
Tidskriftsartikel (refereegranskat)abstract
- This paper presents a picoliter sample preparation technique utilizing the flow-through principle, allowing on-line coupling of chromatographic systems to be made. The work was performed in order to investigate the characteristics and the physicochemical properties of the sample preparation using typical mobile phase conditions from μ-CLC (column liquid chromatography) separations. The device presented here is a pressure pulse- driven dispenser, formed by two silicon structures processed by conventional micromachining. The pressure pulse is generated in the flow-through channel by a piezoceramic element. Depending on the orifice size, the droplets ejected range between 30 and 200 pL. The maximum ejection frequency is 500 Hz, limited by resonances within the unit. A pyramid-shaped nozzle improves the directivity of the droplets since it reduces the wetting of the orifice front surface area. The risk of particles sticking close to the orifice is also minimized. The analyses of the deposited sample spots were carried out on a matrix-assisted laser desorption/ionization time-of-flight mass spectrometer with delayed extraction. It was possible to detect attomole amounts (159-248 amol) of various proteins (cytochrome c, ribonuclease A, lysozyme, and myoglobin) from a single droplet of matrix:analyte 1:1 (drop volume ≃ 110 pL). Additionally, it was found that sample enrichment could be carried out using multiple depositions on the same spot; i.e., 31 nM of insulin was easily detected when more than four depositions were made on the same spot, while no detection was possible without sample enrichment. Size optimization of the MALDI sample spot gave target zones of 100-500-μm diameter that matched the size of the laser focal point and resulted in a considerably increased sample throughput.
|
|
| 10. |
- Önnerfjord, Patrik, et al.
(författare)
-
Tyrosinase graphite-epoxy based composite electrodes for detection of phenols
- 1995
-
Ingår i: Biosensors and Bioelectronics. - 0956-5663. ; 10:6-7, s. 607-619
-
Tidskriftsartikel (refereegranskat)abstract
- The characterization and analytical performance of a tyrosinase graphite-epoxy electrode for the detection of phenolic compounds are described. The biocomposite configuration is based on the entrapment of commercially available tyrosinase in a graphite-epoxy matrix, and the mixing of the resulting conductive epoxy resin with a hardener. The enzyme electrode is mounted as a working electrode in an amperometric flow cell of the confined wall-jet type and studied in the flow injection mode. The bioprobe is electrochemically characterized by hydrodynamic and cyclic voltammetry for catechol and phenol. An applied potential of -100 mV vs. Ag/AgCl is found to be optimal for electrochemical reduction of the enzyme products (quinone forms) for the biocomposite electrode. The dependence of the response of the biocomposite on the flow rate, the amount of loaded enzyme, the buffer composition, pH, and oxygen is investigated. The response of the biosensor to different phenolic compounds is also evaluated. The limits of detection (S/N = 3) for phenol and catechol were 1·0 μM and 0·04 μM, respectively. No loss in response could be detected after 100 injections of catechol (R.S.D. <2%). Stability of the biocomposite depends on storage conditions. Theoretical advantages described in the literature for biocomposite electrodes, for example, repolishing and bulk modification, are empirically studied in this work.
|
|
