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PubMed1986

A semiautomated analysis method for catecholamines, indoleamines, and some prominent metabolites in microdissected regions of the nervous system: an isocratic HPLC technique employing coulometric detection and minimal sample preparation.

The application of a commercially available coulometric electrochemical detector to the automated HPLC analysis of some monoamines and their metabolites in microdissected areas of the rat nervous system is described. Apart from the stability and high sensitivity of the system, other appealing features of the technique are the facile sample preparation and long-term sample storage characteristics which show minimal analyte degradation. Basal values of some regional monoamine and metabolite concentration are listed together with a brief appendix that serves as a user's guide to the operation and maintenance of the detection system.

3,4-Dihydroxyphenylacetic Acid↗
PubMed2003

Sample preparation effects in matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry of partially depolymerised carboxymethyl cellulose.

Sample preparation effects in matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOFMS) of partially depolymerised carboxymethyl cellulose (CMC) have been investigated. The depolymerisation was either enzymatic or acidic. Fractions of enzymatically depolymerised CMC were collected from size-exclusion chromatography (SEC) and further investigated by MALDI-TOFMS. 2,5-Dihydroxybenzoic acid was used as matrix, dissolved in H(2)O due to the poor solubility of CMC in suitable organic solvents. The samples were dried by two methods, in ambient atmosphere and at reduced pressure. Under reduced pressure the sample spot homogeneity increased. This drying method, however, produced additional adduct peaks in the mass spectra originating from ion exchange on the CMC oligomers. Analysis of CMC could be performed in both negative and positive ion modes. Mass discrimination and variation in ionisation efficiency were demonstrated by comparing mass spectra with SEC data. Measurements of the degree of substitution (DS) were performed on three CMCs with different DS values, which were depolymerised in trifluoroacetic acid. The three CMCs were easily distinguished from one another, but the obtained DS values deviated from the values supplied by the manufacturer.

Artifacts↗
PubMed2004

Liquid-phase microextraction utilising plant oils as intermediate extraction medium--towards elimination of synthetic organic solvents in sample preparation.

Hollow fibre based liquid-phase microextraction (LPME) using fatty oils and essential oils as the organic phase was evaluated to develop sample preparation technology eliminating the use of hazardous organic solvents. Basic drugs were extracted from different aqueous samples (0.2 to 1 mL) through approximately 15 microL of either almond oil, arachis oil, olive oil, soy-bean oil, anise oil, fennel oil, lavender oil, or peppermint oil (organic phase) immobilised within the pores of a polypropylene hollow fibre and into 20 microL of 10 mM HCOOH (acceptor phase) present inside the lumen of the hollow fibre. The extraction performance of the essential oils was comparable with the solvents normally used in LPME (dihexyl ether, n-octanol, and dodecyl acetate) in terms of extraction recovery and extraction speed. Whereas all essential oils tested were compatible with human urine, only anise oil was successful for plasma. The fatty oils provided lower recoveries than the essential oils due to higher viscosity, but all the fatty oils were compatible both with urine and plasma samples. In spite of the multi-component nature of the oils tested, they were not found to seriously contaminate the acceptor phases during extraction. In conclusion, fatty oils and essential oils may serve as alternative organic phase in LPME, eliminating the use of hazardous organic solvents.

Mentha piperita↗
PubMed1980

Comparison of three sample preparation techniques for determination of organic bromide and chloride in halogenated lipids by x-ray fluorescence spectroscopy.

Three sample preparation techniques--thin-film, solution, and cellulose pellet--were applied to the determination of bromide in brominated lipids by X-ray fluorescence spectroscopy. Using brominated vegetable oils of known bromide content it was demonstrated that the thin-film technique could result in erroneously high bromide contents, which could also vary with the amount of oil applied, depending on the solvent used. As solutions in hexane, slightly high bromide contents were observed at high concentrations. With the cellulose pellets, bromide contents similar to known values were observed. It was concluded that the cellulose pellet procedure, although more time consuming, and less convenient for ready recovery of sample, was the most suitable for organic bromide determination. Similar results were indicated for chlorinated oils.

Fats↗
PubMed1997

Variation in weight of stool samples prepared by the Kato-Katz method and its implications.

We investigated both the extent of the variation in weight of stool samples prepared by the Kato-Katz method and its influenced on egg counts and commonly used group parameters of infection derived from them. In a first study group of 795 people, the total mean weight of stool aliquots, prepared with templates designed to contain 28.3 mm3, was 23.0 mg with 95% of the individual values lying between 12.0 and 34.0 mg. Minimum and maximum values were 2.4 and 49.5 mg, respectively. Frequency distributions of the individual weights, in series of slides prepared by different laboratory assistants, showed significant differences. In a second study group of 199 people, duplicate series of slides were prepared and variations in the weight of examined stool were related to variations in egg count. The correlation between repeated individual sample weights in this series was poor, but the correlation between eggs counts was good. This was translated, at aggregate level, in very similar classifications in egg count categories. This classification was also hardly influenced by the choice of the conversion factor to transform eggs counts per slide into eggs per gram. At the individual level, the variability in egg counts far outweighed the variability in sample weight and was not clearly related to it. We therefore concluded that variations in weight of examined stool are considerable, but account for only a minimal part of the important egg count fluctuations generally observed.

Adult↗
PubMed2006

Sample preparation for precise and quantitative electron holographic analysis of semiconductor devices.

Wedge polishing was used to prepare one-dimensional Si n-p junction and Si p-channel metal-oxide-silicon field effect transistor (pMOSFET) samples for precise and quantitative electrostatic potential analysis using off-axis electron holography. To avoid artifacts associated with ion milling, cloth polishing with 0.02-microm colloidal silica suspension was used for final thinning. Uniform thickness and no significant charging were observed by electron holography analysis for samples prepared entirely by this method. The effect of sample thickness was investigated and the minimum thickness for reliable results was found to be approximately 160 nm. Below this thickness, measured phase changes were smaller than expected. For the pMOSFET sample, quantitative analysis of two-dimensional electrostatic potential distribution showed that the metallurgical gate length (separation between two extension junctions) was approximately 54 nm, whereas the actual gate length was measured to be approximately 70 nm by conventional transmission electron microscopy. Thus, source and drain junction encroachment under the gate was 16 nm.

Journal Article↗
PubMed1986

Robotic sample preparation evaluated for the immunochemical determination of cardiac isoenzymes.

A general-purpose bench-top laboratory robot was programmed to perform sample preparation for immunoprecipitative and immunoinhibitory determinations of cardiac isoenzymes with commercial kits. Run size could be varied from 1 to 24 patients' samples, and was determined by the robot without prompting. The robot placed the processed samples into a sample rotor for dried transfer to a batch centrifugal analyzer. Robotic precision compared well with that of manual performance of the same procedures at four concentrations of lactate dehydrogenase (EC 1.1.1.27) isoenzyme 1 activity and at one of two concentrations of creatine kinase (EC 2.7.3.2.) isoenzyme MB activity; for the other creatine kinase MB concentration, use of the robot significantly improved precision. Results for 100 samples from patients after open-heart surgery were highly correlated with manually obtained results for both isoenzyme determinations (r less that 0.96 each). Time to completion for small run sizes for either approach was comparable about 25 min. For larger batches, robotic run time increased rapidly, to 169 min for 24 patients' samples vs 41 min by manual assay.

Autoanalysis↗
PubMed1986

[Radioimmunoassay of the progestagen dienogest using various methods of sample preparation of plasma].

For the radioimmunological determination (RIA) of the progestagen dienogest (1, 17 alpha-Cyanomethyl-17 beta-hydroxy-estra-4,9-dien-3-one) in plasma three methods of sample preparation were tested and compared: Extraction of plasma samples with dichlormethane (I), binding of plasma dienogest to an antiserum added; removal of non-bound steroids by means of activated charcoal, extraction of dienogest using dichlormethane (II), using the RIA without extraction of plasma samples after partial precipitation of proteins by means of ammonium sulphate (III). The reliability of the dienogest-RIA is, characterized by a limit of detection of 3.2 pg (I, III) and 5 pg (II) per tube, respectively, by "within-assay"- and "between-assay" variation coefficients of 3 to 5% and 3 to 9%, respectively, in parallel determinations and by a high rate of recovery of dienogest (greater than 90%) added to plasma. The application of the parallelism test method to different plasma volumes confirms the accuracy of method I and II. When method III was applied to plasma samples with low concentrations of dienogest parallelism wasn't found in all cases.

Chromatography, High Pressure Liquid↗
PubMed1994

PCR based detection of Mycobacterium tuberculosis: effect of sample preparation.

Tests based on the polymerase chain reaction (PCR) for the detection of the Mycobacterium tuberculosis complex in clinical samples have a lower sensitivity when compared to culture. This has been attributed to the presence of inhibitors to Taq polymerase and/or suboptimal DNA extraction procedures. We tested different methods of processing smear negative culture positive sputum (n = 52) using different detergents, including nonidet P-40 (NP-40), sodium dodecyl sulphate (SDS), tween 20, triton X 100 and N-lauryl sarcosine. The detergents were used in combination with lysozyme and proteinase K enzymes. NP-40 was significantly better than SDS, tween 20 and N lauryl sarcosine (p < 0.05). When NP-40 was used as the detergent, 42 out of 52 specimens gave positive results with the standard amplification protocol which amplifies a 245 bp sequence of the insertion element IS 986. The 10 specimens that were negative were further diluted ten fold and/or eluted in sephadex G-50 columns before standard DNA amplification. A further 8 specimens then became positive. Elution in sephadex G-50 was better than ten fold dilution in processing of samples. The two negative samples had very low colony counts (n < 5). The study demonstrates that the sensitivity of the PCR is dependent on the sample preparation technique and the amount of target sequence available for amplification.

DNA, Bacterial↗
PubMed2002

Collection, storage, and filtration of in vivo study samples using 96-well filter plates to facilitate automated sample preparation and LC/MS/MS analysis.

The benefits of high-throughput bioanalysis within the pharmaceutical industry are well established. One of the most significant bottlenecks in bioanalysis is transferring in vivo-generated study samples from their collection tubes during sample preparation and extraction. In most cases, the plasma samples must be stored frozen prior to analysis, and the freeze/thaw (F/T) process introduces thrombin clots that are capable of plugging pipets and automated liquid-transfer systems. A new approach to dealing with this problem involves the use of Ansys Captiva 96-well 20-microm polypropylene filter plates to collect, store frozen, and filter plasma samples prior to bioanalysis. The samples are collected from the test subjects, and the corresponding plasma samples are placed directly into the wells of the filter plate. Two Duoseal (patent pending) covers are used to seal the top and bottom of the plate, and the plate is stored at down to -70 degrees C. Prior to sample analysis, the seals are removed and the plate is placed in a 96-well SPE manifold. As the plasma thaws, it passes (by gravity or mild vacuum) through the polypropylene filter into a 96-well collection plate. A multichannel pipet or automated liquid-transfer system is used to transfer sample aliquots without fear of plugging. A significant advantage of this approach is that, unlike other methods, issues related to incomplete pipetting are virtually eliminated. The entire process is rapid since thawing and filtering take place simultaneously, and if a second F/T cycle is required for reanalysis, it is not necessary to refilter the samples (additional clotting was not observed after three F/T cycles). This technique was tested using monkey, rat, and dog plasma and sodium heparin and EDTA anticoagulants. To assess the possibility of nonspecific binding to the polypropylene filter, a variety of drug candidates from diverse drug classes were studied. Validation data generated for two Lilly compounds from distinct classes, before and after filtering, are presented in this paper as practical examples of this technique. While LC/MS/MS is the primary method of bioanalysis in our laboratory, the technique presented in this paper is applicable to other forms of detection as well.

Autoanalysis↗
PubMed1986

Rapid sample preparation method for the determination of chloramphenicol in swine muscle by high-performance liquid chromatography.

A simple and rapid sample preparation method for the determination of chloramphenicol in swine muscle tissue at the 10 micrograms/kg level is described. The method comprises sonication-aided extraction with ethyl acetate, addition of hexane to the extract and cleaning up and concentration of the extract on a small column packed with silica gel. Analysis was performed by high-performance liquid chromatography on a ChromSep column with ChromSpher C8 using acetonitrile-sodium acetate buffer as the mobile phase. Detection was performed at 280 nm. Mean recoveries from spiked muscle samples were 79 +/- 3% (10-50 micrograms/kg). The distribution of chloramphenicol in different muscle and fatty tissues from a pig to which a single dose of chloramphenicol was administered was also investigated.

Adipose Tissue↗
PubMed2006

Rapid sample preparation method for LC-MS/MS or GC-MS analysis of acrylamide in various food matrices.

A fast and easy sample preparation procedure for analysis of acrylamide in various food matrices was developed and optimized. In its first step, deuterated acrylamide internal standard is added to 1 g of homogenized sample together with 5 mL of hexane, 10 mL of water, 10 mL of acetonitrile, 4 g of MgSO4, and 0.5 g of NaCl. Water facilitates the extraction of acrylamide; hexane serves for sample defatting; and the salt combination induces separation of water and acetonitrile layers and forces the majority of acrylamide into the acetonitrile layer. After vigorous shaking of the extraction mixture for 1 min and centrifugation, the upper hexane layer is discarded and a 1 mL aliquot of the acetonitrile extract is cleaned up by dispersive solid-phase extraction using 50 mg of primary secondary amine sorbent and 150 mg of anhydrous MgSO4. The final extract is analyzed either by liquid chromatography-tandem mass spectrometry or by gas chromatography-mass spectrometry (in positive chemical ionization mode) using the direct sample introduction technique for rugged large-volume injection.

Acrylamide↗
PubMed1995

Sensitive high-performance liquid chromatographic analysis of amlodipine in human plasma with amperometric detection and a single-step solid-phase sample preparation.

A narrow-bore HPLC assay with electrochemical detection for the determination of the calcium antagonist amlodipine in human plasma samples is presented. By using a single-step solid-phase extraction procedure on Bond Elut C2 columns, the sample preparation step has been considerably simplified and less time-consuming compared to earlier presented works. With a linear and reproducible calibration curve over the range 0.5-20 ng ml-1 plasma, the assay has successfully been used in the analysis of more than 500 plasma samples from a multicenter trial.

Amlodipine↗
PubMed2006

Massively parallel sample preparation for the MALDI MS analyses of tissues.

Investigation of the peptidome of the nervous system containing large, often easily identifiable neurons has greatly benefited from single-cell matrix-assisted laser desorption/ionization (MALDI) mass spectrometry and has led to the discovery of hundreds of novel cell-to-cell signaling peptides. By combining new sample preparation methods and established protocols for bioanalytical mass spectrometry, a high-throughput, small-volume approach is created that allows the study of the peptidome of a variety of nervous systems. Specifically, approximately single-cell-sized samples are rapidly prepared from thin tissue slices by adhering the tissue section to a glass bead array that is anchored to a stretchable membrane. Stretching the membrane fragments the tissue slice into thousands of individual samples, their dimensions predominately governed by the size of the individual glass beads. Application of MALDI matrix, followed by the repeated condensation of liquid microdroplets on the fragmented tissue, allows for maximal analyte extraction and incorporation into MALDI matrix crystals. During extraction, analyte migration between the pieces of tissue on separate beads is prevented by the underlying hydrophobic substrate and by controlling the size of the condensation droplets. The procedure, while general in nature, may be tailored to the needs of a variety of analyses, producing mass spectra equivalent to those acquired from single-cell samples.

Animals↗
PubMed2002

Automated sample preparation by pressurized liquid extraction-solid-phase extraction for the liquid chromatographic-mass spectrometric investigation of polyphenols in the brewing process.

The analysis of polyphenols from solid plant or food samples usually requires laborious sample preparation. The liquid extraction of these compounds from the sample is compromised by apolar matrix interferences, an excess of which has to be eliminated prior to subsequent purification and separation. Applying pressurized liquid extraction to the extraction of polyphenols from hops, the use of different solvents sequentially can partly overcome these problems. Initial extraction with pentane eliminates hydrophobic compounds like hop resins and oils and enables the straightforward automated on-line solid-phase extraction as part of an optimized LC-MS analysis.

Automation↗
PubMed2004

Quantification of parvovirus B19 DNA using COBAS AmpliPrep automated sample preparation and LightCycler real-time PCR.

The COBAS AmpliPrep instrument (Roche Diagnostics GmbH, D-68305 Mannheim, Germany) automates the entire sample preparation process of nucleic acid isolation from serum or plasma for polymerase chain reaction analysis. We report the analytical performance of the LightCycler Parvovirus B19 Quantification Kit (Roche Diagnostics) using nucleic acids isolated with the COBAS AmpliPrep instrument. Nucleic acids were extracted using the Total Nucleic Acid Isolation Kit (Roche Diagnostics) and amplified with the LightCycler Parvovirus B19 Quantification Kit. The kit combination processes 72 samples per 8-hour shift. The lower detection limit is 234 IU/ml at a 95% hit-rate, linear range approximately 10(4)-10(10) IU/ml, and overall precision 16 to 40%. Relative sensitivity and specificity in routine samples from pregnant women are 100% and 93%, respectively. Identification of a persistent parvovirus B19-infected individual by the polymerase chain reaction among 51 anti-parvovirus B19 IgM-negative samples underlines the importance of additional nucleic acid testing in pregnancy and its superiority to serology in identifying the risk of parvovirus B19 transmission via blood or blood products. Combination of the Total Nucleic Acid Isolation Kit on the COBAS AmpliPrep instrument with the LightCycler Parvovirus B19 Quantification Kit provides a reliable and time-saving tool for sensitive and accurate detection of parvovirus B19 DNA.

Antibodies, Viral↗
PubMed2006

Comparison of sample preparation methods combined with fast gas chromatography-mass spectrometry for ultratrace analysis of pesticide residues in baby food.

Four sample preparation techniques were compared for the ultratrace analysis of pesticide residues in baby food: (a) modified Schenck's method based on ACN extraction with SPE cleaning; (b) quick, easy, cheap, effective, rugged, and safe (QuEChERS) method based on ACN extraction and dispersive SPE; (c) modified QuEChERS method which utilizes column-based SPE instead of dispersive SPE; and (d) matrix solid phase dispersion (MSPD). The methods were combined with fast gas chromatographic-mass spectrometric analysis. The effectiveness of clean-up of the final extract was determined by comparison of the chromatograms obtained. Time consumption, laboriousness, demands on glassware and working place, and consumption of chemicals, especially solvents, increase in the following order QuEChERS < modified QuEChERS < MSPD < modified Schenck's method. All methods offer satisfactory analytical characteristics at the concentration levels of 5, 10, and 100 microg/kg in terms of recoveries and repeatability. Recoveries obtained for the modified QuEChERS method were lower than for the original QuEChERS. In general the best LOQs were obtained for the modified Schenck's method. Modified QuEChERS method provides 21-72% better LOQs than the original method.

Child↗