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PubMed2001

Fully automated 96-well liquid-liquid extraction for analysis of biological samples by liquid chromatography with tandem mass spectrometry.

A fully automated high-throughput liquid-liquid extraction (LLE) methodology has been developed for preparation of biological samples using a 96-well LLE plate and a 96-channel robotic liquid handling workstation. The 96-well LLE plate is made of a 96-well filter plate filled with inert diatomaceous earth particles, allowing continuous and efficient extraction of analytes between the aqueous biological sample and the organic extraction solvent. Two carboxylic acid-based protease inhibitor compounds with high and low levels of plasma protein binding were chosen for the development and application of the automated methodology. The LLE extracts of the plasma samples of the two compounds were analyzed by high-performance liquid chromatography with electrospray (ESI) tandem mass spectrometry (LC-MS/MS). The LC-MS/MS method was developed using a rapid gradient LC separation, followed by sample introduction through an ionspray interface in the negative ion mode and tandem mass spectrometric detection with selected reaction monitoring. In the optimized LLE method, a formate buffer solution was first loaded into a 96-well filter plate packed with inert diatomaceous earth material. Then crude plasma samples and a water-immiscible organic solvent, methyl ethyl ketone, were sequentially added to the LLE plate so that LLE would occur in the interface between the two liquid phases on the surface of individual particles in each well. The organic eluate containing extracted analytes was evaporated and reconstituted for LC-MS/MS analysis. This fully automated LLE methodology avoids several disjointed steps involved in a manual or semiautomated LLE method, leading to significantly reduced sample preparation time, increased sample throughput, and clean sample extracts for improved ESI-MS/MS detection. The automated LLE methodology is universal and can be employed for sample preparation of other biological fluids. The complete bioanalytical method, based on the automated LLE and fast gradient LC-MS/MS, was validated and successfully applied to the quantitative analysis of protease inhibitors in rat plasma.

Animals↗
PubMed2003

Calibration for on-site analysis of hydrocarbons in aqueous and gaseous samples using solid-phase microextraction.

Rapid sampling and sample preparation methodology was investigated using adsorptive poly(dimethylsiloxane)/divinylbenzene and Carboxen/poly(dimethylsiloxane) solid-phase microextraction (SPME) fiber coatings and volatile aromatic hydrocarbons (BTEX: benzene, toluene, ethylbenzene, and o-xylene). A flow-through system was used to generate a standard aqueous solution of BTEX as model sample with known linear velocity. Parameters that affect the extraction process, including sampling time, concentration, water velocity, and temperature, were investigated. Very short sampling times from 10 s and sorbents with strong affinity and large capacity were used to ensure the effect of '"zero sink" and to calibrate the extraction process in the initial linear extraction region. Several different concentrations were investigated, and it was found that mass uptake changes with concentration linearly. The increase of water velocity increases mass uptake, though the increase is not linear. Temperature does not affect mass uptake significantly under typical field sampling conditions. To further accurately describe rapid SPME analysis of aqueous samples, a new model translated from heat transfer to a circular cylinder in cross-flow was used. An empirical correlation to this model was used to predict the mass-transfer coefficient. Findings indicate that predicted mass uptake compares well with experimental mass uptake. The new model was tested for rapid air sampling, and it was found that this new model also predicted rapid air sampling accurately. Findings presented in this study extend the existing fundamental knowledge related to rapid sampling/sample preparation with SPME.

Journal Article↗
PubMed2005

High-throughput nanoliter sample introduction microfluidic chip-based flow injection analysis system with gravity-driven flows.

In this work, a simple, robust, and automated microfluidic chip-based FIA system with gravity-driven flows and liquid-core waveguide (LCW) spectrometric detection was developed. The high-throughput sample introduction system was composed of a capillary sampling probe and an array of horizontally positioned microsample vials with a slot fabricated on the bottom of each vial. FI sample loading and injection were performed by linearly moving the array of vials filled alternately with 50-microL samples and carrier, allowing the probe inlet to enter the solutions in the vials through the slots sequentially and the sample and carrier solution to be introduced into the chip driven by gravity. The performance of the system was demonstrated using the complexation of o-phenanthroline with Fe(II) as a model reaction. A 20-mm-long Teflon AF 2400 capillary (50-microm i.d., 375-microm o.d.) was connected to the chip to function as a LCW detection flow cell with a cell volume of 40 nL and effective path length of 1.7 cm. Linear absorbance response was obtained in the range of 1.0-100 microM Fe(II) (r2=0.9967), and a good reproducibility of 0.6% RSD (n=18) was achieved. The sensitivity was comparable with that obtained using conventional FIA systems, which typically consume 10,000-fold more sample. The highest sampling throughput of 1000 h-1 was obtained by using injection times of 0.08 and 3.4 s for sample and carrier solution, respectively, with a sample consumption of only 0.6 nL for each cycle.

Journal Article↗
PubMed1999

pH-mediated field amplification on-column preconcentration of anions in physiological samples for capillary electrophoresis.

Two limitations of capillary electrophoresis (CE) are the low sample loadability of the capillary and an incompatibility with high ionic strength samples. Several strategies have been described to preconcentrate and lower the ionic strength of physiological samples prior to CE analysis. These have included both off-capillary and on-capillary approaches. We have previously described a version of on-column field-amplification stacking termed pH-mediated stacking. pH-mediated stacking was initially developed for the separation of cations. In this report, we describe the application of pH-mediated sample stacking to anions. In this method, an electrokinetic injection is used to introduce analyte anions into the CE system and simultaneously replace the sample matrix cations with ammonia from the background electrolyte. Base is then electrokinetically injected to neutralize the sample zone and create a low conductivity region across which the analyte anions will stack. Using this method, a sensitivity enhancement of more than 66-fold was achieved without loss in separation efficiency relative to normal electrokinetic injection. Detection limits of 0.3 microM for four phenolic acids in a physiological sample were achieved using simple UV absorbance detection. The limit to the amount of sample that could be loaded using this technique was the length of the separation capillary. To further increase the amount of sample that could be loaded, a double-capillary system was developed. Using the double-capillary system the sensitivity was increased more than 300-fold and detection limits of 0.06 microM were achieved.

Ammonia↗
PubMed2003

As, Hg, and Se flue gas sampling in a coal-fired power plant and their fate during coal combustion.

As, Hg, and Se are the most volatile elements in the flue gas from a coal-fired power plant. Significant amounts of these elements cause an undesired direct gaseous emission, which leads to a serious environmental health risk. The main focus of this study is to evaluate the possibility of simultaneous sampling of these volatile elements using an accurate official method for Hg (the most volatile element). A study of As, Hg, and Se emissions from a 1400 MW coal-fired power plant equipped with electrostatic precipitators (ESPs) was carried out for the combustion of a mixture of two types of coal. Simultaneous sampling of coal, bottom ash, fly ash, flue gas, and particles associated with the gas phase has been performed. Flue gas has been sampled by the Ontario Hydro Method Sampling Train, an ASTM method for Hg speciation. This sampling method was tested for As and Se sampling. As and Se determinations have been performed by HG-AAS, and Hg has been determined by CV-AAS. The results were used to examine the following: overall mass balances, relative distribution of these elements in the coal-fired power plant; As, Hg, and Se concentrations in coal and combustion residues; and predominant oxidation state for Hg in flue gas. The mass balances obtained for As, Hg, and Se were satisfactory in all cases; nevertheless, relative enrichment values in fly ash for As and Se were low; therefore, we concluded that As sampling in flue gas can be conducted by application of the Ontario Hydro Method; nevertheless Se released in the gas phase is not completely collected by this sampling train. Application of this sampling method allowed for performance of Hg speciation. The results indicated that Hg(II) was the predominant species in flue gas. It has also been proved that 24%, more than 99.8%, and 90% for As, Hg, and Se in the stack emissions, respectively, were in the gaseous phase.

Air Pollutants↗
PubMed2004

Principal component analysis applied to Fourier transform infrared spectroscopy for the design of calibration sets for glycerol prediction models in wine and for the detection and classification of outlier samples.

Principal component analysis (PCA) was used to identify the main sources of variation in the Fourier transform infrared (FT-IR) spectra of 329 wines of various styles. The FT-IR spectra were gathered using a specialized WineScan instrument. The main sources of variation included the reducing sugar and alcohol content of the samples, as well as the stage of fermentation and the maturation period of the wines. The implications of the variation between the different wine styles for the design of calibration models with accurate predictive abilities were investigated using glycerol calibration in wine as a model system. PCA enabled the identification and interpretation of samples that were poorly predicted by the calibration models, as well as the detection of individual samples in the sample set that had atypical spectra (i.e., outlier samples). The Soft Independent Modeling of Class Analogy (SIMCA) approach was used to establish a model for the classification of the outlier samples. A glycerol calibration for wine was developed (reducing sugar content < 30 g/L, alcohol > 8% v/v) with satisfactory predictive ability (SEP = 0.40 g/L). The RPD value (ratio of the standard deviation of the data to the standard error of prediction) was 5.6, indicating that the calibration is suitable for quantification purposes. A calibration for glycerol in special late harvest and noble late harvest wines (RS 31-147 g/L, alcohol > 11.6% v/v) with a prediction error SECV = 0.65 g/L, was also established. This study yielded an analytical strategy that combined the careful design of calibration sets with measures that facilitated the early detection and interpretation of poorly predicted samples and outlier samples in a sample set. The strategy provided a powerful means of quality control, which is necessary for the generation of accurate prediction data and therefore for the successful implementation of FT-IR in the routine analytical laboratory.

Calibration↗
PubMed2004

Sensitive PCR analysis of animal tissue samples for fragments of endogenous and transgenic plant DNA.

An optimized DNA extraction protocol for animal tissues coupled with sensitive PCR methods was used to determine whether trace levels of feed-derived DNA fragments, plant and/or transgenic, are detectable in animal tissue samples including dairy milk and samples of muscle (meat) from chickens, swine, and beef steers. Assays were developed to detect DNA fragments of both the high copy number chloroplast-encoded maize rubisco gene (rbcL) and single copy nuclear-encoded transgenic elements (p35S and a MON 810-specific gene fragment). The specificities of the two rbcL PCR assays and two transgenic DNA PCR assays were established by testing against a range of conventional plant species and genetically modified maize crops. The sensitivities of the two rbcL PCR assays (resulting in 173 and 500 bp amplicons) were similar, detecting as little as 0.08 and 0.02 genomic equivalents, respectively. The sensitivities of the p35S and MON 810 PCR assays were approximately 5 and 10 genomic equivalents for 123 bp and 149 bp amplicons, respectively, which were considerably less than the sensitivity of the rbcL assays in terms of plant cell equivalents, but approximately similar when the higher numbers of copies of the chloroplast genome per cell are taken into account. The 173 bp rbcL assay detected the target plant chloroplast DNA fragment in 5%, 15%, and 53% of the muscle samples from beef steers, broiler chickens, and swine, respectively, and in 86% of the milk samples from dairy cows. Reanalysis of new aliquots of 31 of the pork samples that were positive in the 173 bp rbcL PCR showed that 58% of these samples were reproducibly positive in this same PCR assay. The 500 bp rbcL assay detected DNA fragments in 43% of the swine muscle samples and 79% of the milk samples. By comparison, no statistically significant detections of transgenic DNA fragments by the p35S PCR assay occurred with any of these animal tissue samples.

Animal Feed↗
PubMedDate not supplied

Development and evaluation of a micro- and nanoscale proteomic sample preparation method.

Challenges associated with the efficient and effective preparation of micro- and nanoscale (micro- and nanogram) clinical specimens for proteomic applications include the unmitigated sample losses that occur during the processing steps. Herein, we describe a simple "single-tube" preparation protocol appropriate for small proteomic samples using the organic cosolvent, trifluoroethanol (TFE) that circumvents the loss of sample by facilitating both protein extraction and protein denaturation without requiring a separate cleanup step. The performance of the TFE-based method was initially evaluated by comparisons to traditional detergent-based methods on relatively large scale sample processing using human breast cancer cells and mouse brain tissue. The results demonstrated that the TFE-based protocol provided comparable results to the traditional detergent-based protocols for larger, conventionally sized proteomic samples (>100 microg protein content), based on both sample recovery and numbers of peptide/protein identifications. The effectiveness of this protocol for micro- and nanoscale sample processing was then evaluated for the extraction of proteins/peptides and shown effective for small mouse brain tissue samples (approximately 30 microg total protein content) and also for samples of approximately 5000 MCF-7 human breast cancer cells (approximately 500 ng total protein content), where the detergent-based methods were ineffective due to losses during cleanup and transfer steps.

Animals↗
PubMed1999

Do plasma concentrations obtained from early arterial blood sampling improve pharmacokinetic/pharmacodynamic modeling?

In pharmacokinetic/pharmacodynamic (PK/PD) modeling the first blood sample is usually taken 1 to 2 min after drug administration (late sampling). Therefore, investigators have to extrapolate the plasma concentration to Time 0. Extrapolation, however, erroneously assumes instantaneous and complete mixing of drug in the central volume of distribution. We investigated whether plasma concentrations obtained from early arterial blood sampling would improve PK/PD modeling. In 14 pigs, one of five neuromuscular blocking agents (NMBAs) was administered into the right ventricle within 1 sec and arterial sampling was performed every 1.2 sec (1st min). The response of the tibialis muscle was measured mechanomyographically. The influence of inclusion of data from early arterial sampling on PK/PD modeling was determined. Furthermore, the concentrations in the effect compartment at 50% block (EC50) derived from modeling were compared to the measured concentration in plasma during a steady state 50% block. A very high peak in arterial plasma concentration was seen within 20 sec after administration of the NMBA. Extensive modeling revealed that plasma concentrations obtained from early arterial blood sampling improve PK/PD modeling. Independent of the type of modeling, the EC50 and KeO based on data sets that include early arterial blood sampling were, for all five NMBAs, significantly higher and lower respectively, than those based on data sets obtained from late sampling. Early arterial sampling shows that the mixing of the NMBA in the central volume of distribution is incomplete. A parametric PD (sigmoid Emax) model could not describe the time course of effect of the NMBAs adequately.

Animals↗
PubMed2001

Detection and molecular typing of Borrelia burgdorferi sensu lato in Ixodes ricinus ticks and in different patient samples from southwest Germany.

The prevalence of different genospecies of Borrelia burgdorferi sensu lato in infected ticks could be a determinant for the risk of acquiring Lyme borreliosis (LB) and its clinical presentation. A total of 7373 ticks and 2761 samples from LB patients from the same area in southwest Germany were analyzed by PCR to assess the frequency of the occurrence of LB-associated genospecies. Fifteen percent of the tick samples and 19% of the human samples were found positive for the presence of B. burgdorferi sensu lato. Further identification of 1106 B. burgdorferi sensu lato positive tick samples by reverse line blotting and 125 positive patient samples by nested PCR using species-specific primers revealed the occurrence of B. afzelii, B. burgdorferi sensu stricto, B. garinii and B. valaisiana. Both single-species and mixed infections were noted and a similar distribution of the different genospecies was found in ticks compared with human samples. It was also the purpose of this study to obtain more information about a possible correlation between the distribution of Borrelia species and clinical syndromes of LB. Skin biopsies of 59 patients with acrodermatitis chronica atrophicans and cerebrospinal fluid samples from 78 patients with possible neuroborreliosis were analyzed. In conclusion, the distribution of the different genospecies in ticks is the decisive factor for the occurrence of the different Borrelia genospecies in samples from LB patients. Borrelia afzelii is the predominant genospecies in all kind of samples from the observed area and there seems to be no association of particular Borrelia genospecies with distinct clinical manifestations of LB.

Animals↗
PubMed2004

Vitality of oligozoospermic semen samples is improved by both swim-up and density gradient centrifugation before cryopreservation.

PURPOSE: To ascertain whether washing sperm from oligozoospermic and normozoospermic samples before cryopreservation improves post-thaw vitality. METHODS: Normozoospermic (n = 18) and oligozoospermic (n = 16) samples were divided into three aliquots. The first aliquot remained untreated and the second and third aliquots were subjected to the swim-up and discontinuous density gradient sperm washing techniques respectively. Vitality staining was performed, samples mixed with cryopreservation media and frozen. Spermatozoa were thawed, stained, and vitality quantified and expressed as the percentage of live spermatozoa present. RESULTS: Post-thaw vitality in untreated aliquots from normozoospermic samples (24.9% +/- 2.3; mean +/- SEM) was significantly higher (unpaired t-tests; P < 0.01) than untreated oligozoospermic samples (11.9% +/- 2.3). Post-thaw vitality was significantly higher after swim-up in normozoospermic samples (35.6% +/- 2.1; P < 0.001; one-way ANOVA) and oligozoospermic samples (27.7% +/- 1.7; P < 0.01). Density gradient centrifugation significantly improved post-thaw vitality in oligozoospermic (22.4% +/- 1.0; P < 0.01) but not normozoospermic (30.8% +/- 1.8) samples. CONCLUSIONS: Sperm vitality in cryopreserved oligozoospermic samples was improved by both the swim-up and density gradient centrifugation washing techniques prior to freezing.

Cell Separation↗
PubMed2000

Influence of sampling interval on the evaluation of nocturnal blood pressure in subjects with and without obstructive sleep apnoea.

Blood pressure (BP) variability during sleep is high in obstructive sleep apnoea syndrome (OSAS). How BP sampling interval affects the estimate of mean nocturnal BP in OSAS and control subjects was investigated. Nine subjects with apnoea/hypopnoea index (AHI) <5 and 18 OSAS patients with AHI >30 underwent nocturnal polysomnography with beat-by-beat BP monitoring. Mean nocturnal BP was evaluated averaging: a) all systolic (Ps) and diastolic (Pd) BP values; b) Ps and Pd sampled every 5, 10, 15, 20, and 30 min. The sampling starting point was repeatedly shifted, and several mean BP estimates for each sampling interval were obtained. Differences (deltaPs and deltaPd) between means obtained by sampling BP and by averaging all BP values were calculated. In both groups deltaPs and deltaPd scatter increased as sampling interval increased; their variance was always higher in OSAS subjects (p<0.001). Over 95% of deltaPs and deltaPd were <5% of the beat-by-beat mean values at all sampling intervals in controls, but this occurred only at sampling intervals < or =10 min in OSAS subjects. To conclude, for each blood pressure sampling time, a larger number of inaccurate nocturnal mean blood pressure estimates are obtained in obstructive sleep apnoea syndrome than in control subjects. Obstructive sleep apnoea syndrome subjects require more frequent blood pressure measurements to obtain a similar accuracy in nocturnal blood pressure evaluation.

Blood Pressure↗
PubMed1999

Sampling of periodontal pathogens by paper points: evaluation of basic parameters.

Paper points are widely established for the collection of subgingival plaque or other oral samples to analyze the microbiota, especially the presence of periodontal pathogenic bacteria such as Actinobacillus actinomycetemcomitans or Porphyromonas gingivalis. In contrast to the high frequency of usage of paper points in oral sampling, very few data are available about the parameters influencing the sampling process. Therefore, in four different in vitro experiments (6-9 repeats), we inoculated paper points with standardized suspensions (2 x 10(9) colony-forming units/ml) of A. actinomycetemcomitans and P. gingivalis to test the influence of the origin (kind) of paper point ("manufacturer"), size (according to the International Organization for Standardization ISO 25-80), sampling time (5 to 60 s), and elution time (5 to 60 s). Sampled bacteria were detected and (semi-) quantified using 16S rRNA/DNA-directed oligonucleotide probes. The bacterial load was categorized and calculations performed with index values ranging between 0 (below the detection limit of 10(2) to 10(3) colony-forming units) and 9 (> 10(6) colony-forming units). We found statistically significant differences in the efficiency of bacterial sampling between the 5 paper point manufacturers tested, expressed in a mean bacterial index between 4.4 and 7.8. ISO 45 Paper points were statistically proven to work most efficiently. According to our results, a sampling time of 60 s seems to be optimum. However, shorter times between 5 and 30 s did not significantly reduce the sampling efficiency. We found an interval of 20 s best to elute bacteria from the paper points by vortexing. The evaluation of basic parameters for subgingival plaque sampling by paper points might help to optimize microbiologically based diagnostics in periodontal diseases.

Aggregatibacter actinomycetemcomitans↗
PubMed2000

Comparison of zinc reduction with platinum reduction for analysis of deuterium-enriched water samples for the doubly labeled water technique.

OBJECTIVE: Isotope ratio mass spectrometry of hydrogen and oxygen is frequently used to determine total energy expenditure (TEE) using doubly labeled water. Conventionally, hydrogen isotope ratio is determined in hydrogen gas generated from water samples using zinc reduction. We compare this with a new automated platinum method to determine the ratios of hydrogen isotopes in deuterium-enriched water samples. RESEARCH METHODS AND PROCEDURES: The platinum method of sample preparation was compared with the zinc method in three ways: analytical variation in deuterium enrichment (within sample; n = 51), analytical variation in TEE estimates (within sample set; n = 10), and level of agreement of TEE estimates between both methods (n = 14). RESULTS: For the zinc method, the standard deviation for multiple sets of triplicate 2H2O sample analysis was +/-4.36 per thousand and +/-2.07 per thousand for platinum. The correlation between TEE estimates when sample sets were analyzed in duplicate was r = 0.89 for zinc and r = 0.83 for platinum. The intercept and slope of the regression line were significantly different from the line of identity for duplicate TEE estimates by zinc but were not different from the line of identity for platinum. After correction for the intra-assay variation of each method, the correlation between zinc and platinum for TEE was 0.77, and the intercept, but not the slope, of the regression was significantly different from the line of identity. The mean difference between the zinc method and the platinum method was 56 kcal/day, and the 95% confidence interval was -438 to 550 kcal/day. DISCUSSION: These data suggest that the platinum method is at least as reliable as the zinc method as a sample preparation technique for isotope ratio mass spectrometry of deuterium-enriched water samples. The platinum method is also less costly and less labor-intensive than the zinc method.

Autoanalysis↗
PubMed2002

Sampling private wells at past homes to estimate arsenic exposure: a methodologic study in New England.

We are conducting a collaborative, population-based case-control study in Maine, New Hampshire, and Vermont to investigate the reasons for the elevated bladder cancer mortality in northern New England. Arsenic in drinking water is one of the primary exposures under investigation. To estimate subjects' lifetime exposure to waterborne arsenic, it will be necessary to obtain water samples from private wells that subjects used in the past. We conducted a methodologic study to assess the feasibility of locating and sampling from private wells at subjects' past residences. Ninety-eight New Hampshire residents (mean age 67 years) completed a questionnaire requesting the complete address, dates of occupancy, and drinking water sources for each home lived in since birth. An interviewer then asked subjects for more detailed information about each home to assist in a field search of past homes in the three-state study area of Maine, New Hampshire, and Vermont. Fifty-eight of the 98 subjects indicated that they had used a total of 103 private wells in 95 previous homes located in these three states. We conducted a field search to locate these 95 homes, visited town offices to find the properties on tax maps and obtain the current owners' names and addresses, attempted to obtain permission from the current owners to sample the wells, and collected water samples. In all, 48 (47%) of the 103 past wells in the study area were sampled successfully. The remaining wells were not sampled because the homes were not located (22%) or had been demolished (2%), permission to sample the wells was not obtained (17%), the wells had been destroyed (7%) or could not be found on the grounds of the residence (3%), or for other reasons (2%). Various approaches for improving the success rates for sampling water from private wells are discussed, as is the use of predictive modeling to impute exposures when sampling is not feasible.

Adult↗
PubMed2005

Comparative analysis of the effects of sample source and test methodology on the assessment of protein expression in acute myelogenous leukemia.

Numerous studies have analyzed the expression and prognostic importance of various proteins in acute myelogenous leukemia (AML). We sought to determine whether the sample source and methodology used to measure protein expression affect the results obtained. To determine the importance of sample source, we used Western blotting to compare the expression of eight proteins and phosphoproteins in the leukemia blast-enriched fraction of 118 blood- and 108 marrow-derived samples, including 37 paired samples. To determine the importance of methodology, the expression of five proteins was measured in 20 paired samples by Western blotting, laser scanning cytometry (LSC), and flow cytometry. The mean expression and range of expression in blood- and marrow-derived samples were statistically identical for all eight proteins. Expression measurements for the 37 paired blood and marrow samples also had very high statistical correlation. The LSC and flow cytometry data had the highest concordance when compared using Kolmogorov-Smirnoff D-stats (range of R values, 0.8-1.0). High concordance was also observed between the LSC and flow cytometry results when the percentage of cells positive for expression was dichotomized into positive or negative expression. However, there was less correlation between LSC and flow cytometry when the actual percentages of positive cells were compared. The majority of discordant situations involved samples that were positive by flow cytometry but negative by LSC. The correlation between Western blotting signal intensity and the percentage of expression-positive cells measured by LSC or flow cytometry varied by protein but was limited when there was little heterogeneity in expression by either method. In conclusion, provided that leukemia blast-enriched fractions were analyzed, the blood- and marrow-derived samples had identical protein expression. There was good concordance of results between flow cytometry and LSC, which share similar technology, but more limited correlation between these methods and Western blotting.

Blotting, Western↗
PubMed1997

Sampling intercomparisons for aldehydes in simulated workplace air.

Thirty one laboratories of various EU Member States have participated in two interlaboratory comparisons in order to assess errors of personal sampling methods associated with both the sampling and the analytical steps. In contrast to conventional quality control schemes, this project particularly focuses attention on the sampling and identification step; it is executed by means of sampling exercises and has included discussions on potential sources of error. In a sampling exercise, participants come to a central facility and perform measurements on synthetic workplace air in a laboratory installation. Concentration levels of formaldehyde, acrolein, glutaraldehyde and acetaldehyde between 0.1 and 2 times the limit value for workplace air were prepared at various humidity levels and with acetone, occasionally, as interferent. Sampling times varied from 1-4 h. The related analytical work is performed at the analyst's own laboratory. The intention is for each participant to determine the observed value of the delivered standard atmosphere using the sampling method of his own choice. Trueness (bias), precision and relative overall uncertainty of each method-laboratory combination is calculated and verified towards compliance with EN 482, which outlines minimum performance criteria. The first challenge involved the precise gas phase generation of the selected analytes in high air flows (up to 300 1 min-1) and calculating the true value only by direct reference to primary standards. This was accomplished by modifying the capillary dosage injection technique so that reactive compounds, like low molecular mass aldehydes, could be dosed with the same accuracy and precision as unreactive solvents. A permeation tube with high emission rate was developed for formaldehyde. Up to ten different sampling techniques were evaluated. The measurement methods used by the majority of the participants were based on pumped sampling on silica cartridges (or tubes) and glass fiber filters, coated with 2,4-dinitrophenylhydrazine. It was observed that for formaldehyde, and in some cases for acetaldehyde and glutaraldehyde, the majority of the method-laboratory combinations complied with an overall uncertainty of 30%. The results for acrolein, however, indicated a systematic negative bias, often larger than minus 50% of the true value, caused by the decomposition of the acrolein DNPH derivative in the presence of excess acid and excess DNPH.

Air Pollutants, Occupational↗
PubMed1999

Evaluation of two adsorbents for diffusive sampling and thermal desorption-gas chromatographic analysis of monoterpenes in air.

Tube type samplers with two different adsorbents, Chromosorb 106 and Tenax TA, were evaluated by laboratory experiments and field tests for simultaneous diffusive sampling of alpha-pinene, beta-pinene and delta 3-carene and subsequent thermal desorption-gas chromatographic analysis. No statistically significant effects of exposure time, concentrations of monoterpenes or relative humidity were found for samplers with Chromosorb 106 when running a factorial design, with the exception of the adsorption of delta 3-carene, for which some weak effects were noted. Samplers with Tenax TA were affected by the sampling time as well as the concentration for all terpenes, with a strong interaction effect between these two factors. The terpenes showed good storage stability on both adsorbents. No effect of back-diffusion was noted when using Chromosorb 106, while Tenax TA showed some back-diffusion effects. The uptake rates, in ml min-1, for the terpenes on Chromosorb 106 were 0.36 for alpha-pinene, 0.36 for beta-pinene and 0.40 for delta 3-carene. The corresponding average values on Tenax TA were 0.30 for alpha-pinene, 0.32 for beta-pinene and 0.38 for delta 3-carene. The field validation proved that diffusive sampling on Chromosorb 106 agreed well with pumped sampling on charcoal for stationary samples, while the personal samples indicated a discrepancy of 25% between Chromosorb 106 and charcoal samples. Tenax TA generally gave lower results than Chromosorb 106 in all field samples. Samplers packed with Chromosorb 106 could be used to monitor terpene levels in workplaces such as sawmills. The major advantages with this method are the sampling procedure, which is simple to perform compared to other techniques, the easily automated analysis procedure and the possibility to reuse the samplers.

Adsorption↗