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PubMed1998

Limited-sample NMR using solenoidal microcoils, perfluorocarbon plugs, and capillary spinning.

This study demonstrates three improvements to mass-limited NMR using solenoidal microcoils as detectors: (1) sample confinement using liquid perfluorocarbon plugs to increase the observe factor, (2) design and incorporation of a capillary spinner to improve spectral line widths, and (3) facile sample changing via the use of a capillary insert. The probe is constructed to spin a fused silica capillary of 530 microns i.d., 700 microns o.d. inside a solenoidal coil wound around a 0.8 mm i.d., 1 mm o.d. glass capillary. The smaller capillary contains the sample, and capillaries with different samples can be exchanged easily. In high-resolution limited sample microcoil NMR studies published thus far, the length of the sample plug has been 7-10 times the length of the solenoid to avoid line broadening from volume magnetic susceptibility (chi v) mismatches at both ends of the sample. This arrangement is not efficient since it places most of the sample volume outside of the coil observe volume. It is shown here that the observe factor cannot exceed 23% if the sample plug is bracketed by air, without substantial line broadening occurring. However, if the sample is bracketed by two liquid perfluorocarbon plugs, the observe factor can be increased to 70% while maintaining high spectral resolution. This is equivalent to improving the limit of detection by a factor of 3, or reducing the total data acquisition time for a given signal-to-noise by a factor of 9. It is also shown that, for the 440-nL sample plug used in this study (bracketed by the perflurocarbon plugs), sample spinning can improve the spectral resolution from 1.5 (nonspinning) to 0.6 Hz (spinning). This corresponds to a further improvement in the limit of detection of 2.5, or just over a factor of 6 decrease in data acquisition time.

Fluorocarbons↗
PubMed2005

An automated electrokinetic continuous sample introduction system for microfluidic chip-based capillary electrophoresis.

An automated and continuous sample introduction system for microfluidic chip-based capillary electrophoresis (CE) was developed in this work. An efficient world-to-chip interface for chip-based CE separation was produced by horizontally connecting a Z-shaped fused silica capillary sampling probe to the sample loading channel of a crossed-channel chip. The sample presentation system was composed of an array of bottom-slotted sample vials filled alternately with samples and working electrolyte, horizontally positioned on a programmable linearly moving platform. On moving the array from one vial to the next, and scanning the probe, which was fixed with a platinum electrode on its tip, through the slots of the vials, a series of samples, each followed by a flow of working electrolyte was continuously introduced electrokinetically from the off-chip vials into the sample loading channel of the chip. The performance of the system was demonstrated in the separation and determination of FITC-labeled arginine and phenylalanine with LIF detection, by continuously introducing a train of different samples. Employing 4.5 kV sampling voltage (1000 V cm(-1) field strength) for 30 s and 1.8 kV separation voltage (400 V cm(-1) field strength) for 70 s, throughputs of 36 h(-1) were achieved with <1.0% carryover and 4.6, 3.2 and 4.0% RSD for arginine, FITC and phenylalanine, respectively (n = 11). Net sample consumption was only 240 nL for each sample.

Electrophoresis, Capillary↗
PubMed2005

Comparison of 24 h averaged VOC monitoring results for residential indoor and outdoor air using Carbopack X-filled diffusive samplers and active sampling--a pilot study.

Analytical results obtained by thermal desorption GC/MS for 24 h diffusive sampling of 11 volatile organic compounds (VOCs) are compared with results of time-averaged active sampling at a known constant flow rate. Air samples were collected with co-located duplicate diffusive sampling tubes and one passivated canister. A total of eight multiple-component sampling events took place at fixed positions inside and outside three private homes. Subsequently, a known amount of sample air was transferred from the canister to an adsorbent tube for analysis by thermal desorption GC/MS. Results for the 11 most prevalent compounds--Freon 11, 1,3-butadiene, benzene, toluene, tetrachloroethene, ethylbenzene, m,p-xylene, o-xylene, 4-ethyltoluene, 1,3,5-trimethylbenzene, and p-dichlorobenzene--show that the ratio of average study values (diffusive sampling to active sampling) is 0.92 with 0.70 and 1.14 extreme ratios. Absolute percent difference for duplicate samples using diffusive sampling was <10% for the four most prevalent compounds. Agreement between the two sampling approaches indicates that the prediction of approximately constant diffusive sampling rates based on previous laboratory studies is valid under the field conditions.

Air Pollutants↗
PubMed2000

Influence of sexual stimulation on sperm parameters in semen samples collected via masturbation from normozoospermic men or cryptozoospermic men participating in an assisted reproduction programme.

To evaluate the influence of sexual stimulation via sexually stimulating videotaped visual images (VIM) on sperm function, two semen samples were collected from each of 19 normozoospermic men via masturbation with VIM. Two additional samples were collected from each man via masturbation without VIM. The volume of seminal plasma, total sperm count, sperm motility, percentage of morphologically normal spermatozoa, outcome of hypo-osmotic swelling test and zona-free hamster oocyte sperm penetration assay, and markers of the secretory function of prostate were significantly larger in semen samples collected via masturbation with VIM than masturbation without VIM. The improved sperm parameters in the samples collected via masturbation with VIM may reflect an enhanced prostatic secretory function and increased loading of the vas deferens at that time. In a similar protocol, two semen samples were collected via masturbation with VIM from each of 22 non-obstructed azoospermic men. Semen samples from these men had been occasionally positive in the past for a very small number of spermatozoa (cryptozoospermic men). Two additional samples were collected from each cryptozoospermic man via masturbation without VIM. The volume of seminal plasma, total sperm count, sperm motility, and a marker of the secretory function of prostate were significantly larger in semen samples collected via masturbation with VIM. Fourteen out of the 22 men were negative for spermatozoa in both samples collected via masturbation without VIM. These men demonstrated spermatozoa in both samples collected via masturbation with VIM. Six men with immotile spermatozoa in both samples collected via masturbation without VIM exposed motile spermatozoa in both samples collected via masturbation with VIM. High sexual stimulation during masturbation with VIM results in recovery of spermatozoa of greater fertilizing potential both in normozoospermic and cryptozoospermic men. The appearance of spermatozoa after masturbation with VIM in the vast majority of cryptozoospermic men is of clinical significance in programmes applying intracytoplasmic sperm injections for the management of severe male infertility and obviates the need for testicular biopsy.

Biomarkers↗
PubMed2001

Iohexol as a marker of glomerular filtration rate in patients with diabetes: comparison of multiple and simplified sampling protocols.

AIMS: To report on the reproducibility of iohexol glomerular filtration rate (GFR) estimation, to compare the plasma clearance of iohexol with that of[51Cr]EDTA and to evaluate the reliability of reduced sampling schedules in estimating GFR in Type 1 and Type 2 diabetes mellitus. METHODS: Agreement was assessed in 15 Type 1 and 26 Type 2 diabetics with creatinine ranging from 53 to 564 micromol/l. RESULTS: The regression between multiple-sample iohexol and[51Cr]EDTA clearances was 0.999 in Type 1 and 0.987 in Type 2 diabetes (P < 0.0001 for both). A seven-sample design and the three-sample approach by Brøchner-Mortensen were validated by comparison with the full-sample schedule in 87 patients (51 Type 1, 36 Type 2). Full-sample GFR was 80.3 +/- 43.8, seven-sample 79.5 +/- 43.9 (r = 0.990) and three-sample 79.8 +/- 45.2 ml.min-1.1.73 m-2 (r = 0.972). The coefficients of variation of GFR were 2.7 +/- 1.4% and 3.8 +/- 1.9% for the full-sample and the seven-sample approaches, respectively, and significantly higher for the three-sample design (6.9 +/- 3.4%, P = 0.0001). CONCLUSIONS: After iohexol injection, the Brøchner-Mortensen schedule does not provide an accurate estimate of GFR. The seven-sample approach gives acceptable errors and allows a good estimate of GFR throughout a wide range of renal function.

Adult↗
PubMed2001

Quantitation of genomic DNA in plasma and serum samples: higher concentrations of genomic DNA found in serum than in plasma.

BACKGROUND: Plasma and serum samples have been used to detect cell-free genomic DNA in serum or plasma in certain pathologic conditions such as systemic lupus erythematosus, pulmonary embolism, and malignancies, as well as in fetal cell chimerisms in maternal serum and/or plasma. In this study, baseline concentrations of cell-free DNA in serum and plasma samples were evaluated for the study of posttransfusion chimerism. STUDY DESIGN AND METHODS: DNA was extracted from fresh or stored (4 degrees C for 1-6 days) normal donor serum or plasma samples (ACD; EDTA) by using reagents from an HIV assay kit. After incubation and washing of samples, purified DNA was amplified with HLA DQ-alpha primers (GH26 and 27) or human Y-chromosome primers (SA and SD) to quantitate the concentration of genomic DNA. RESULTS: Fresh serum samples had concentrations of cell-free DNA that were about 20-fold higher than the concentrations in fresh plasma samples. The concentration of cell-free genomic DNA in serum samples increased daily, to a level more than 100 times baseline after clotted blood tubes were stored at 4 degrees C for 4 to 5 days. There was a small increase in cell-free plasma DNA in stored ACD whole blood samples. Male WBCs, spiked into fresh nonanticoagulated female blood, were lysed during the process of clotting, with male DNA liberated into the serum samples. CONCLUSION: Most cell-free DNA in serum samples is generated during the process of clotting in the original collection tube. The concentration of cell-free genomic DNA in fresh plasma is probably the same as that in circulation. Consequently, while serum samples should not be used to monitor the concentration of cell-free DNA in a patient's circulation, serum collected from sample tubes containing clots (i.e., without anticoagulant), 3 to 5 days after the date of phlebotomy, could be useful as a source of DNA with which to screen for posttransfusion microchimerism.

Blood Donors↗
PubMed2001

Comparison of the sensitivity of NAT using pooled donor samples for HBV and that of a serologic HBsAg assay.

BACKGROUND: Studies were conducted using samples from early and late-stage HBV-infected persons to determine the pool size at which PCR had better sensitivity than a sensitive HBsAg chemoluminescence immunoassay (CLIA-HBsAg). STUDY DESIGN AND METHODS: HBV seroconversion panels were tested for HBsAg by CLIA and for HBV DNA by nested PCR (95% hit rate: 100 copies/mL); PCR was carried out at various dilutions. HBV serologically positive samples that were detected from the simultaneous screening of 540,161 routine whole-blood donations using CLIA-HBsAg and agglutination assays were also characterized for additional markers of HBV infection. RESULTS: In 9 of 10 HBV seroconversion panels, PCR had better sensitivity than CLIA-HBsAg at dilutions of 1-in-25 or lower. Of 65 CLIA-only confirmed-positive donor samples (agglutination assay-negative), 8 represented early infection, 2 of which were PCR positive at a 1-in-50 dilution but negative at a 1-in-100 dilution. Only 2 of 47 samples from probable late-stage HBV infection that were positive on CLIA only were PCR positive with 0.1-mL sample volume and the S-region primer; the remaining 45 samples required a 1.0-mL sample input and C-region primer for increased PCR positivity. The remaining 10 CLIA-only confirmed-positive donor samples were from HBV vaccine recipients. None of the 12 CLIA- and HBsAg-negative donor samples that were strongly anti-HBc reactive could be detected by PCR at any dilution; all 12 were PCR positive when undiluted, but 4 required a 1.0-mL input volume for PCR positivity. CONCLUSION: For the detection of samples representing early-stage HBV infection, PCR at dilutions of 1-in-25 or lower (equivalent to a pool of < or =25 members) had greater sensitivity than CLIA-HBsAg. In contrast, samples from late-stage HBV infection were detected by PCR only with undiluted samples (0.1-mL or 1.0-mL input volumes), regardless of CLIA-HBsAg reactivity. Therefore, although NAT using minipools of 25 donations or less may be effective for the detection of early-stage HBV infection, it may not be effective for the detection of persistent HBV infection.

Blood↗
PubMed2006

Effect of ambient storage of wheat samples on their nutritive value for chickens.

1. Two wheat cultivars (Abbott and Equinox) were grown using the same crop husbandry conditions. Batches of each cultivar were stored at ambient temperature (AT) for 0, 6, 12, 18 or 24 weeks. Those stored at AT for less than 24 weeks were then stored at -20 degrees C up to the end of the storage period. The 24 week (AT) samples were divided into two lots and one lot was frozen to -20 degrees C for 24 h. 2. Each of the 12 wheat samples was included (650 g/kg) in a nutritionally complete diet and fed to broiler chickens from 7 to 21 d of age. The apparent metabolisable energy (AME) of each sample was determined using a rapid bioassay with adult cockerels. 3. The Equinox cultivar sample had a high proportion (92/1000) of visibly sprouted grains and had a low Hagberg falling number (high amylase activity) and high content of free sugars compared to the Abbott sample. There were no major changes in dry matter, free sugar content, water extract viscosity or Hagberg falling number due to the different times of ambient storage of either of the two wheat cultivar samples. 4. The chickens fed on the Abbott cultivar wheat had a greater weight gain, although there were no differences in the determined AME of the two wheat cultivars. 5. The Abbott sample stored at ambient temperature for 6 weeks had a greater AME than the sample given no ambient storage and the samples given 18 and 24 weeks ambient storage. There were no significant differences between the Equinox cultivar samples. There were no effects of storage time on the growth performance of the broiler chickens. 6. The AME of the unfrozen Abbott cultivar was greater than the frozen (for 24 h) sample but there was no effect on broiler growth performance. This study suggests that, in some wheat samples, there may be a benefit in avoiding freezing during storage.

Animal Feed↗
PubMed2000

Stability of YKL-40 concentration in blood samples.

The stability of YKL-40, a mammalian member of the family of 18 glycosylhydrolases, in blood samples handled under different temperatures and different time intervals before centrifugation was studied in paired serum and plasma samples from 25 healthy premenopausal Danish women. Significant elevations of YKL-40 were found in 8 paired serum samples left on the clot for more than 3 h at room temperature compared to paired serum samples left on the clot for 3 h or less. Significant elevations of YKL-40 were found in 8 paired plasma (EDTA) samples left on the blood cells for more than 8 h at room temperature compared to paired plasma (EDTA) samples left on the blood cells for 8 h or less. No elevations were found in YKL-40 levels in serum samples left on the clot at 4 degrees C for 24 h or in plasma (EDTA) samples left on the blood cells for 72 h before centrifugation. Significantly lower concentrations of YKL-40 were measured in plasma (EDTA) compared with paired serum samples with a serum/plasma ratio of 1.4 in samples left on the clot or on blood cells at 4 degrees C for up to 24 h. Repetitive freezing and thawing had no significant effect on the measured YKL-40 concentrations. In conclusion, we have shown that YKL-40 is very dependent on the handling procedures. All the blood samples must be processed into plasma (EDTA) within 8 h at room temperature or into serum in less than 3 h at room temperature. If this is not possible, the blood samples must be stored at 4 degrees C until processed.

Adipokines↗
PubMed2000

Determination of total sulfur compounds and benzothiazole in asphalt fume samples by gas chromatography with sulfur chemiluminescence detection.

As part of a collaborative project between the National Institute for Occupational Safety and Health and the Federal Highway Administration to evaluate asphalt pavers' exposures to asphalt fume and their potential health effects, a method was developed for the determination of total sulfur compounds and benzothiazole in asphalt fume samples. Asphalt fume samples were collected from asphalt mixtures with and without the addition of ground-up rubber tires. The asphalt fume samples were collected with sampling trains that consisted of a Teflon membrane filter and an XAD-2 adsorbent tube. Filter and sampling tube media were extracted with hexane and subsequently analyzed by gas chromatography with a sulfur chemiluminescence detector. Separation was achieved with a 100 percent dimethyl polysiloxane fused silica column. Typical calibration curves had linear correlation coefficients of 0.99 or better with a relative standard deviation (RSD) of 5 percent. Benzothiazole desorption efficiency (DE) determined using spiked sampling tubes ranged from 96.5 percent at 5.0 micrograms to 89.4 percent at 40 micrograms with RSD values from 0.9 to 4.0 percent. Benzothiazole storage recovery determined using sampling tubes spiked at 20 micrograms and refrigerated for 30 days at 4 degrees C was 89.8 percent when corrected for the DE with an RSD of 1.1 percent. The limit of detection for the method determined using spiked sampling tubes was 0.30 microgram. Quantitation for total sulfur compounds and benzothiazole was against benzothiazole standards in hexane. Because of detector selectivity, sample preparation consisted of a simple hexane extraction even when samples had a high background due to hydrocarbon overload. Detector sensitivity provided quantitation in the sub-microgram region. Because of the sample preparation step and because benzothiazole was determined during the same analysis run, this method is straightforward and analytically efficient. The method has been used to analyze asphalt fume samples collected at several asphalt paving and roof operations.

Air Pollutants, Occupational↗
PubMed1990

Electrostatic effects in asbestos sampling. I: Experimental measurements.

Electrostatic charge can cause errors during sampling of airborne asbestos fibers and other particles. The change in particle trajectories caused by charge effects during sampling can result in nonuniform deposits on the collecting filter surface and net loss of sample. The degree of these electrostatic effects depends on particle charge, sampler charge, sampler conductivity, and sampling flow rate and direction. The purpose of this research was to evaluate the dependence of sampling efficiency and sample uniformity on these variables. Humidity has been postulated as a primary determinant of particle charge during aerosol generation. Measurements of particle charge and concentration were made as a function of relative humidity with chrysotile fibers generated from a fluidized bed. A strong increase in charge and a decrease in concentration of fibers was noted as the relative humidity was decreased below 15%. The effects of conductive versus nonconductive samplers and sampling flow rate were measured as a function of particle and sampler charge levels. Nonconductive samplers can carry a large and variable charge distribution on their surfaces. This can result in a biased and highly variable particle deposit on the filter when sampling charged particles. Conductive cowls spread any acquired charge over the entire surface and produce a more symmetrical and less biased charged particle deposit. Increasing the sampling flow rate will improve sampling efficiency and decrease deposit variability because the charged particle has less time to interact with the field produced by the sampler. These results suggest that sampling problems caused by electrostatic charge interactions are most likely to occur under low humidity conditions of dust generation, that sampling should be done at as high a flow rate as possible to reduce these effects, and that analysts should select fields toward the center of the filter to minimize bias and variability.

Aerosols↗
PubMed2004

Comparison of the Microgenics CEDIA heroin metabolite (6-AM) and the Roche Abuscreen ONLINE opiate immunoassays for the detection of heroin use in forensic urine samples.

Current Department of Defense (DoD) and Department of Health and Human Services (HHS) procedures for the detection of heroin abuse by testing urine utilize an initial opiate (codeine/morphine) immunoassay (IA) screen followed by gas chromatography-mass spectrometry (GC-MS) confirmation of 6-acetylmorphine (6-AM), if the morphine concentration is above established cutoff. An alternative to the current opiates screen for heroin abuse is the direct IA for the metabolite of heroin, 6-acetylmorphine. In this regard, the performance of the Microgenics CEDIA heroin metabolite (6-AM) screening reagent was assessed. This evaluation was conducted on the P module of a Hitachi Modular automated IA analyzer calibrated using 6-AM at 10 ng/mL. Reproducibility, linearity, accuracy, sensitivity, and interferences associated with use of the 6-AM IA reagent were evaluated. The IA reagent precision (percent coefficient of variation (%CV)) around each of seven standards was less than 0.63%, with a linearity (r(2)) value of 0.9951. A total of 37,713 active duty service members' urine samples were analyzed simultaneously using the CEDIA heroin metabolite (6-AM) reagent and the Roche Abuscreen ONLINE opiate reagent to evaluate both the prevalence rate of 6-AM in the demographic group and the sensitivity and specificity of the reagents for the detection of heroin use. Of the 37,713 samples tested using the CEDIA heroin metabolite (6-AM) reagent, three samples screened positive at the DoD and HHS cutoff of 10 ng/mL. One of the three samples confirmed positive for 6-AM by GC-MS above the cutoff of 10 ng/mL, the two remaining samples confirmed negative for 6-AM at a GC-MS limit of detection (LOD) of 2.1 ng/mL. In contrast, the Roche Abuscreen ONLINE opiate IA produced 74 opiate-positive results for codeine/morphine, with 6 of the 74 specimens confirming positive for morphine above the DoD cutoff concentration of 4000 ng/mL (8% DoD morphine confirmation rate), only one of the 74 opiate-positive screen specimens confirmed positive for 6-AM above the 10 ng/mL GC-MS cutoff concentration. As a further check of the sensitivity and specificity of the Microgenics 6-AM IA reagent, human urine samples (n = 87) known to contain 6-AM by GC-MS, were re-analyzed using both IA reagents. All 87 of the samples screened positive using the CEDIA heroin metabolite (6-AM) assay. However, using the Roche ONLINE opiate reagent, 12 of the known 6-AM positives screened negative at the DoD and HHS screening cutoff of 2000 ng/mL (morphine). Of the remaining 75 samples that screened positive by the ONLINE opiate reagent, five of the samples did not contain morphine above the DoD GC-MS cutoff concentration of 4000 ng/mL and would not have required 6-AM analysis. However, under the HHS GC-MS morphine cutoff concentration of 2000 ng/mL all 75 samples would have required 6-AM analysis. Furthermore, using the current DoD opiate screen, 17 out of 87 samples known to contain 6-AM would have gone undetected (19.5% false-negative rate); additionally, even under the more stringent HHS opiate screening standards 12 out of the 87 samples known to contain 6-AM would also have gone undetected (13.8% false-negative rate). The Microgenics CEDIA heroin metabolite (6-AM) reagent assay appears well adapted for the rapid and specific detection of heroin abuse as an alternative for, or an adjunct test to, the current opiates (codeine/morphine) IA screening procedure.

Calibration↗
PubMed2005

Use of secondary enrichment for isolation of Salmonella from naturally contaminated environmental samples.

Since the implementation of Hazard Analysis Critical Control Point (HACCP), the need for on-farm food safety risk assessment and management has greatly increased. In order to provide accurate risk assessments, attention should be focused on better characterization of the Salmonella isolation and identification techniques. In this work, we compared the isolation ability of 4 Salmonella-specific protocols: immunomagnetic separation (DB), tetrathionate (TT) broth, Rappaport-Vassiliadis R10 (RV) broth, and a secondary enrichment (TR) procedure as well as 2 selective solid media (brilliant green agar, BG; and xylose-lysine tergitol 4, XLT4). All 4 methods were compared in litter and drag swab samples that were collected weekly during the broiler grow out period in 7 houses. There were 65/126 (51.6%) pooled litter samples positive and 115/304 (37.8%) drag swab samples positive for Salmonella by at least one method. Of the 65 positive litter samples, DB, RV, and TT isolated 1 (2.7%), 31 (47.7%), and 23 (35.4%) of the samples as positive when using BG agar, respectively. The TR protocol identified 83.1% (54/65) of the positive samples as positive when using BG agar. In the drag swab samples, DB did not identify any samples as positive, whereas TT and RV found 28 (25.7%) and 26 (23.9%) of the 109 samples to be positive when using BG agar, respectively. Again, the TR protocol identified the highest percentage of positive samples (94.5%). An analysis of agreement, kappa, revealed that TT and RV did not always agree on which samples were positive, although the number of samples identified as positive by both were not different. A comparison between the 2 agar plates used, BG and XLT4, showed that they had high agreement when the secondary enrichment protocol was used, but agreement was only moderate to low when the other 3 methods were used.

Animals↗
PubMed2003

Genotoxicity of samples of nickel refinery dust.

At the International Nickel Company (INCO) nickel refinery in Clydach, Wales, U.K., which has operated since 1901, 365 respiratory cancers, including 85 nasal cancers and 280 lung cancers, have occurred in workers since the 1920s. From 1901 to 1923, incidences of these cancers were high. In 1923, the refining process was changed, eliminating a nickel arsenide, Ni5As2, called orcelite, from the refinery. Incidences of respiratory cancers decreased substantially from 1925 to 1930. Refinery dust samples were obtained in 1920 and in 1929; both of these samples contain primarily nickel oxide (NiO), but the 1920 sample also contains orcelite. The orcelite content of the 1920 sample is approximately 10%, while that of the 1929 sample is approximately 1%. We hypothesized that orcelite in the 1920 sample was partially responsible for inducing nasal and lung cancers in the refinery workers, and we tested this hypothesis. The 1920 and 1929 samples and orcelite were phagocytosed by cultured C3H/10T1/2 Cl 8 (10T1/2) mouse embryo cells to similar extents and were similarly cytotoxic to 10T1/2 cells. The 1920 sample and orcelite induced dose-dependent morphological transformation of 10T1/2 cells; the 1929 sample did not. The cell transforming ability of the 1920 sample, and therefore its probable carcinogenicity, correlates with induction of respiratory cancers in refinery workers exposed to orcelite-containing nickel refinery dust before 1923. Inability of the 1929 sample to induce morphological transformation correlates with decreased human respiratory tumor incidence at this plant after 1923. This data supports our hypothesis that orcelite in the 1920 refinery sample contributed to its carcinogenicity to nickel refinery workers.

Animals↗
PubMed2002

Tissue microarray sampling strategy for prostate cancer biomarker analysis.

High-density tissue microarrays (TMA) are useful for profiling protein expression in a large number of samples but their use for clinical biomarker studies may be limited in heterogeneous tumors like prostate cancer. In this study, the optimization and validation of a tumor sampling strategy for a prostate cancer outcomes TMA is performed. Prostate cancer proliferation determined by Ki-67 immunohistochemistry was tested. Ten replicate measurements of proliferation using digital image analysis (CAS200, Bacus Labs, Lombard, IL, USA) were made on 10 regions of prostate cancer from a standard glass slide. Five matching tissue microarray sample cores (0.6 mm diameter) were sampled from each of the 10 regions in the parallel study. A bootstrap resampling analysis was used to statistically simulate all possible permutations of TMA sample number per region or sample. Statistical analysis compared TMA samples with Ki-67 expression in standard pathology immunohistochemistry slides. The optimal sampling for TMA cores was reached at 3 as fewer TMA samples significantly increased Ki-67 variability and a larger number did not significantly improve accuracy. To validate these results, a prostate cancer outcomes tissue microarray containing 10 replicate tumor samples from 88 cases was constructed. Similar to the initial study, 1 to 10 randomly selected cores were used to evaluate the Ki-67 expression for each case, computing the 90th percentile of the expression from all samples used in each model. Using this value, a Cox proportional hazards analysis was performed to determine predictors of time until prostate-specific antigen (PSA) recurrence after radical prostatectomy for clinically localized prostate cancer. Examination of multiple models demonstrated that 4 cores was optimal. Using a model with 4 cores, a Cox regression model demonstrated that Ki-67 expression, preoperative PSA, and surgical margin status predicted time to PSA recurrence with hazard ratios of 1.49 (95% confidence interval [CI] 1.01-2.20, p = 0.047), 2.36 (95% CI 1.15-4.85, p = 0.020), and 9.04 (95% CI 2.42-33.81, p = 0.001), respectively. Models with 3 cores to determine Ki-67 expression were also found to predict outcome. In summary, 3 cores were required to optimally represent Ki-67 expression with respect to the standard tumor slide. Three to 4 cores gave the optimal predictive value in a prostate cancer outcomes array. Sampling strategies with fewer than 3 cores may not accurately represent tumor protein expression. Conversely, more than 4 cores will not add significant information. This prostate cancer outcomes array should be useful in evaluating other putative prostate cancer biomarkers.

Biomarkers↗
PubMed1997

Evaluating laboratory usage in the intensive care unit: patient and institutional characteristics that influence frequency of blood sampling.

OBJECTIVES: To develop a predictive equation to estimate the frequency of blood drawing for intensive care unit (ICU) laboratory tests and to evaluate variations in ICU blood sampling practices after adjusting for patient and institutional factors. DESIGN: Prospective, inception, cohort study. SETTING: Forty-two ICUs in 40 hospitals, including 20 teaching and 17 nonteaching ICUs. PATIENTS: A consecutive sample of 17,440 ICU admissions, in which 14,043 blood samples were drawn for laboratory testing on ICU days 2 to 7. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Patient demographic, physiologic, and treatment data were obtained on ICU day 1; the type and number of blood samples for laboratory testing were recorded on ICU days 1 to 7. In the 42 ICUs, a mean of 16.2 blood samples were drawn for tests on ICU days 2 to 7, but varied between 23 samples in the teaching ICUs and 9.9 samples in nonteaching ICUs. Using only ICU day 1 patient data, we predicted the subsequent number of samples drawn on ICU day 2 (R2 = .26 across individual patients) and on ICU days 2 to 7 (R2 = .26 across individual patients). The most important determinants of the number of blood samples drawn on ICU days 2 to 7 were the ICU day 1 Acute Physiology Score and admission diagnosis. After controlling for patient variables, hospital teaching status, number of beds, and location in the East and South were significantly (p < .05) associated with increased blood sampling on ICU day 2 and on ICU days 2 to 7. More frequent use of an arterial cannula and mechanical ventilation were also associated with increased blood sampling on subsequent days. CONCLUSIONS: The ability to adjust for patient and institutional variables and to predict the number of blood samples drawn for laboratory tests can allow ICUs to compare their practices with those of other units. When integrated into a continuous quality improvement process, this information can be used to identify and focus on opportunities for improving blood conservation and reducing excessive diagnostic testing.

APACHE↗
PubMedDate not supplied

Energy concentration of human milk estimated from 24-h pools and various abbreviated sampling schemes.

The concentration of gross energy in 24-h pooled samples of milk is compared with that of single samples obtained during the same period. Samples were collected from mothers exclusively breastfeeding their infants on demand. The objective of the comparison was to evaluate simplified sampling schemes for approximating the concentration of energy of milk produced over 24 h. The lowest concentration of energy of single samples occurred in milk collected between 12 midnight and 6 a.m. The concentration of energy of milk was related negatively to the volume expressed and the time interval preceding each expression. The interval preceding each expression was correlated positively to the volume expressed. These results indicate that the pattern of milk expression is important in the design of sampling schemes. Therefore, sampling schemes may not be transferable between populations with distinct feeding patterns. The highest correlation between single samples and the 24-h value was obtained from samples collected between 12 midnight and 6 a.m. (r = 0.84, p less than 0.001). The widths of the 95% confidence and prediction intervals for the equation relating values from single samples to the 24-h pools were +/- 2.7% and +/- 10.5%, respectively, of the mean 24-h pool sample value when three single samples (collected between 12 midnight and 6 a.m., 6 a.m. and 12 noon, and 6 p.m. and 12 midnight) were included in the predictive equation.

Analysis of Variance↗
PubMed1995

Efficient sampling strategies for forecasting pharmacokinetic parameters of irinotecan (CPT-11): implication for area under the concentration-time curve monitoring.

A linear two-compartment Bayesian pharmacokinetic model was developed using a standard two-stage population method for the novel anti-cancer agent CPT-11 from 11 adult patients with refractory cancer. The accuracy and efficiency of this Bayesian model for estimating pharmacokinetic parameters including the area under the concentration-time curve (AUC) was then evaluated using two different sampling strategies in a new study cohort of 13 patients with cancer. Sampling strategies included either one, two, or three nonsteady-state feedback levels determined empirically and from optimal sampling theory (D-optimality). All 24 patients in this study received CPT-11 (60 mg/m2) as a 90-min infusion. Pharmacokinetic parameters derived from the Bayesian model combined with these limited sampling strategies were compared with those parameters obtained from the full sample data sets (n = 10) analyzed by weighted nonlinear least squares regression (reference method). The least-bias and most precise sampling times for estimating AUC were 3.5; 3.5 and 9.5; and 0.5, 3.5, and 9.5 h, respectively. At these times, only marginal improvement in precision of the AUC estimate was observed using two versus three samples. However, the precision of the estimate of clearance was not improved using two versus three samples. The sampling times derived from optimal sampling theory were 0.25, 3.5, 8.5, and 24 h and correlated closely to the actual and best empirical sampling times of 0.5, 3.5, 9.5, and 24 h. These results strongly suggest that Bayesian estimation combined with only two optimally timed samples accurately predicts the AUC of CPT-11 and should be useful for implementing adaptive control dosing for monitoring CPT-11 systemic exposure in patients with cancer.

Adult↗