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[A digital radiology method for assessing vertebral osteoporosis].

The radiologic identification of vertebral fractures is usually subjective and reproducibility is poor. This paper describes a new digital radiologic method to perform vertebral morphometry, i.e. osteoradiometry (ORM). Lateral radiographs of the thoracic and lumbar spine were obtained in 50 premenopausal women and digitalized by means of a video camera. A special computer software enables to calculate the anterior (Ha), middle (Hm), and posterior (Hp) heights of vertebral bodies (T4-L5) and the morphometric indices of vertebral fractures. ORM reproducibility was assessed by comparing repeated measurements made by two radiologists: the intra- and interobserver variation coefficients (CV) were respectively 1.5% and 2.3% for Hp; 1.3% and 2% for Hm; 1.4% and 2.1% for Ha. The normal range for vertebral dimensions was therefore established. The anterior and posterior heights increased from T4 to L2, but for L3-L5 the posterior height was lower than the anterior height (Ha/Hp > 1). Vertebral heights positively correlated with the standing heights of the subjects (r = 0.2, p < 0.05). Weight and the body mass index (BMI) were not correlated with vertebral heights. These normal values, compared with those found in osteoporosis patients, will allow to assess ORM diagnostic efficacy in identifying vertebral fractures.

Absorptiometry, Photon↗

Determination of SNP allele frequencies in pooled DNAs by primer extension genotyping and denaturing high-performance liquid chromatography.

By testing DNA pools rather than single samples the number of tests for a case-control association study can be decreased to only two for each marker: one on the patient and one on the control pool. A fundamental requirement is that each pool represents the frequency of the markers in the corresponding population beyond the influence of experimental errors. Consequently the latter must be carefully determined. To this aim, we prepared pools of different size (49-402 individuals) with accurately quantified DNAs, estimated the allelic frequencies in the pools of two SNPs by primer extension genotyping followed by DHPLC analysis and compared them with the real frequencies determined in the single samples. Our data show that (1) the method is highly reproducible: the standard deviation of repeated determinations was +/-0.014; (2) the experimental error (i.e., the discrepancy between the estimated and real frequencies) was +/-0.013 (95% C.I.: 0.0098-0.0165). The magnitude of this error was not correlated to the pool size or to the type of SNP. The effect of the observed experimental error on the power of the association test was evaluated. We conclude that this method constitutes an efficient tool for high-throughput association screenings provided that the experimental error is low. We therefore recommend that before a pool is used for extensive association studies, its quality, i.e., the experimental error, is verified by determining the difference between estimated and real frequencies for at least one marker.

Alleles↗

Intraobserver reproducibility of Doppler measurements of uterine artery blood flow velocity in premenopausal women.

OBJECTIVE: To determine the intraobserver repeatability of Doppler measurements of uterine artery blood flow velocity and the contribution of various factors to within-subject variance. DESIGN: Seventeen healthy premenopausal women underwent vaginal Doppler ultrasound examination of the uterine artery by the same observer. Three measurements were taken at each of three sites: 1) the currently recommended sampling site; 2) the ascending branch of the uterine artery at a level between the lower and middle third of the corpus uteri; 3) 1.5 cm lateral to the recommended sampling site. Three measurements were taken at each site. For each measurement, three uniform consecutive cardiac cycles were analyzed. Peak systolic velocity, time-averaged maximum velocity, and pulsatility index were calculated. Each Doppler shift spectrum was analyzed twice. Thus, for each women, 18 measurement results per sampling site were obtained. Analysis of variance was used. RESULTS: The effect of sampling site on measurements of peak systolic velocity and time-averaged maximum velocity was non-significant, but pulsatility index values obtained at the distal sampling site were slightly higher than those obtained at the other sites (P = 0.01). Repetition accounted for most of the within-subject variance. Averaging the results of the three repeat measurements yielded increased intraclass correlation coefficients: 0.79-0.89 for peak systolic velocity, 0.80-0.92 for time-averaged maximum velocity and 0.86-0.93 for pulsatility index. CONCLUSION: As the effect of repetition on the results of Doppler measurements of uterine artery blood flow velocity is large, the average of several repeat measurements should be used to enhance measurement reproducibility. However, it is not worth doing more than one analysis of a Doppler shift spectrum, and it is not worth analyzing more than one cardiac cycle per spectrum.

Adult↗

Reproducibility of a symptomatic response to upright tilt in young patients with unexplained syncope.

Serial upright tilt testing has been advocated as a possible therapeutic end point in treating patients with recurrent syncope, but the reproducibility of such testing has not been well-established in the absence of therapy. In 21 patients with recurrent syncope and a symptomatic response to upright tilt testing, tilt was repeated following a 25- to 30-minute recovery. Syncope or presyncope was reproduced in 14 of 21 patients on repeat tilt; 4 of 21 patients experienced milder symptoms, and 3 of 21 patients remained entirely asymptomatic on repeat tilt. The pattern of physiologic response was different during initial and repeat tilt in 7 of the 21 patients. Blood pressure and heart rate (before and during symptoms) were similar for the group on initial and repeat tilt. The abnormal physiologic response and associated symptoms elicited during upright tilt testing for unexplained syncope can be reproduced on immediate repeat tilt testing in most patients. However, the frequently attenuated response during repeat testing may limit its use as a therapeutic end point in individual patients.

Adolescent↗

Measurement of the margins of partial-coverage tooth preparations for CAD/CAM.

PURPOSE: This study tested the hypothesis that a scanning laser 3-dimensional digitizer is a precise and accurate instrument to measure chamfered and beveled margins of partial coverage tooth preparations for computer-aided design/computer-aided manufacturing (CAD/CAM). MATERIALS AND METHODS: The margins were measured by the digitizer on stone dies and calculated by triangulation into a 3-D representation. Instrument precision was defined as the ability to reproduce the same margin in repeated measurements and expressed as the coefficient of variation as a percentage. Instrument accuracy for chamfered and beveled margins was estimated by correlating their measurements to the measurement of the margin of a spherical calibration "phantom" with known dimensions. Accuracy was expressed as the standard deviation. RESULTS: The precision errors for the box- and cusp-chamfered margins and cusp-beveled margins were 3.9%, 3.4%, and 2.4%, respectively. With regard to accuracy the standard deviations of the measurements of the box- and cusp-chamfered margins and cusp-beveled margins were 19 microns, 21 microns, and 24 microns, respectively, compared to 15 microns for the phantom. CONCLUSION: Measurements of chamfered and beveled margins by a scanning laser 3-D digitizer for CAD/CAM are (1) precise (error < 4%) and (2) accurate, with a standard deviation of less than 9 microns compared to optimal measurements of the spherical margin of the phantom.

Analog-Digital Conversion↗

Cholecystokinin-Stimulated mebrofenin (99mTc-Choletec) hepatobiliary scintigraphy in asymptomatic postcholecystectomy individuals: assessment of specificity, interobserver reliability, and reproducibility.

OBJECTIVES: Cholecystokinin-stimulated hepatobiliary scintigraphy (CCK-HBS) is a noninvasive method reported to be highly accurate in the diagnosis of sphincter of Oddi dysfunction. Our primary aim was to assess the specificity of CCK-HBS by evaluating its ability to exclude disease in 20 asymptomatic postcholecystectomy individuals. Secondary aims were to assess the interobserver reliability in scoring the CCK-HBS examinations between three blinded observers and to assess reproducibility of CCK-HBS repeated in the same individuals. METHODS: Twenty asymptomatic postcholecystectomy individuals with normal liver serum chemistries underwent CCK-HBS on two separate occasions. Three nuclear medicine specialists read each CCK-HBS study in a blinded fashion. RESULTS: There was good agreement between the three observers reading the same scans for both the first scan (kappa = 0.554) and the second scan (kappa = 0.507). There was poor agreement between the first and second scans on the same patient, read by the same nuclear medicine specialist (kappa = 0.062-0.385). The overall specificity of the CCK-HBS score was 77.5%; however, the specificity was only 60% when a true negative was defined as two negative CCK-HBS examinations. CONCLUSIONS: Quantitative CCK-HBS is of poor specificity in asymptomatic postcholecystectomy individuals. Hence, it is of questionable value in excluding sphincter of Oddi dysfunction in patients suspected to suffer from this disorder.

Adult↗

Discriminatory power and reproducibility of novel DNA typing methods for Mycobacterium tuberculosis complex strains.

In recent years various novel DNA typing methods have been developed which are faster and easier to perform than the current internationally standardized IS6110 restriction fragment length polymorphism typing method. However, there has been no overview of the utility of these novel typing methods, and it is largely unknown how they compare to previously published methods. In this study, the discriminative power and reproducibility of nine recently described PCR-based typing methods for Mycobacterium tuberculosis were investigated using the strain collection of the interlaboratory study of Kremer et al. This strain collection contains 90 M. tuberculosis complex and 10 non-M. tuberculosis complex mycobacterial strains, as well as 31 duplicated DNA samples to assess reproducibility. The highest reproducibility was found with variable numbers of tandem repeat typing using mycobacterial interspersed repetitive units (MIRU VNTR) and fast ligation-mediated PCR (FLiP), followed by second-generation spoligotyping, ligation-mediated PCR (LM-PCR), VNTR typing using five repeat loci identified at the Queens University of Belfast (QUB VNTR), and the Amadio speciation PCR. Poor reproducibility was associated with fluorescent amplified fragment length polymorphism typing, which was performed in three different laboratories. The methods were ordered from highest discrimination to lowest by the Hunter-Gaston discriminative index as follows: QUB VNTR typing, MIRU VNTR typing, FLiP, LM-PCR, and spoligotyping. We conclude that both VNTR typing methods and FLiP typing are rapid, highly reliable, and discriminative epidemiological typing methods for M. tuberculosis and that VNTR typing is the epidemiological typing method of choice for the near future.

Bacterial Typing Techniques↗

The role of measurement reliability in clinical trials.

One of the principal characteristics of an outcome measure in a clinical trial, and any measurement in general, is its reliability. Reliability refers to the reproducibility of the measurement when repeated at random in the same subject or specimen. Reliability is often confused with validity, which refers to the extent to which the variable properly measures the underlying trait of interest. The coefficient of reliability is an estimate of the proportion of all variation that is not due to measurement error and is readily estimated from replicate measurements. The reliability of a measurement determines its maximal correlation or R2 and slope (or effect size) in regression models, its sensitivity and specificity when used for classifications or predictions, and the power of a statistical test employing the measurement. All decline as the reliability of the measure declines. The reliability of a measurement is an important consideration in the choice of the primary outcome measure for a clinical trial and in the choice of measures used for assessment of eligibility and exclusion. Reliability of measures should be assessed and assured by a quality control program based on randomly selected duplicate assessments. Just as the power of a study is reported in a final publication, so also should the reliability of the outcome and eligibility measurements so as to allow the authors to better describe, and readers to better understand, the sources of imprecision in study results, and those who follow to improve the design of future trials.

Clinical Trials as Topic↗

Noninvasive measurement of cardiovascular function in mice with high-temporal-resolution small-animal PET.

UNLABELLED: The aim of this study was to explore the feasibility of determining parameters of cardiovascular function in mice noninvasively by high-temporal-resolution imaging with a dedicated small-animal PET system. METHODS: Twenty-five anesthetized mice (28.8 +/- 4.6 g) were injected via an intravenous catheter with a 30-microL bolus of (18)F-FDG (8-44 MBq). The first 9 s of data were reconstructed into 30 frames of 0.3 s using filtered backprojection. The time-activity curve derived from a left ventricle volume of interest was corrected for tracer recirculation and partial volume. Cardiac output was calculated by the Stewart-Hamilton method, in which cardiac output is total injected activity divided by the area under the left ventricle time-activity curve. Cardiac output divided by body weight was defined as cardiac index; cardiac output divided by heart rate yielded the stroke volume. In 5 mice, measurements were repeated 2-4 times to assess reproducibility. In 4 mice, the hemodynamic response to dobutamine was examined by measuring heart rate, cardiac output, and stroke volume. RESULTS: The cardiac output averaged 20.4 +/- 3.4 mL/min; in the repeated measurements, the parameter displayed a mean percentage SD per mouse of 10% +/- 6%. The cardiac index averaged 0.73 +/- 0.19 mL/min/g and the stroke volume 45.0 +/- 6.9 microL, and both correlated with heart rate (r = 0.53, P = 0.007, and r = 0.49, P = 0.01, respectively). During dobutamine stress, heart rate increased from 423 +/- 50 to 603 +/- 30 beats/min (P = 0.002) and cardiac output increased from 18.5 +/- 1.9 to 32.0 +/- 4.2 mL/min (P = 0.008). CONCLUSION: Parameters of cardiovascular function can be measured in mice noninvasively by radionuclide angiography using high-temporal-resolution small-animal PET. Measured values of cardiac output and stroke volume are reproducible and comparable to those obtained with MRI. The approach permits the monitoring of changes in cardiovascular function in response to pharmacologic intervention.

Animals↗

Paraspinal skin temperature patterns: an interexaminer and intraexaminer reliability study.

BACKGROUND: Paraspinal thermography is used by chiropractors as an aid in assessing the presence of vertebral subluxation. Few reliability studies have been carried out, with mixed results. Digital infrared scanning equipment is now available with location tracking that may enhance reproducibility. Digitized scans enable a computer-aided interpretation of thermographic patterns. OBJECTIVE: To assess the ability of examiners to reproduce thermal patterns. STUDY DESIGN: Repeated measures with 2 examiners assessing the same patient on 2 occasions. Thirty asymptomatic students served as subjects. METHODS: A TyTron C-3000 handheld thermographic scanner interfaced to a Microsoft Windows compatible personal computer was used for all recordings. Each examiner recorded 2 scans on each patient. It took an average of 3 minutes to complete all 4 scans. Data were exported to a spreadsheet for initial analysis, then SPSS was used for calculation of intraclass correlation coefficients (ICC). Since the starting and stopping points of scans were not always the same, care was taken to align scans visually, using well-distinguished peaks on the charts as guides. Scans were cropped to remove artifacts that might have occurred at the beginning and end of the scans. Intraexaminer and interexaminer ICCs were calculated. RESULTS: Skin temperatures ranged from 35.4 degrees C to 30.0 degrees C over all scans. The average temperatures changed little from the first to the last scans, indicating that subjects' overall skin temperatures were stable during the scanning procedure. Intraexaminer ICCs ranged from 0.953 to 0.984. The left and right channel data show slightly higher congruence than the Delta channel. The interexaminer reliability coefficients ranged from 0.918 to 0.975. Again, the Delta channel shows slightly less reliability, although the ICCs were quite high for all channels. CONCLUSION: Intraexaminer and interexaminer reliability of paraspinal thermal scans using the TyTron C-3000 were found to be very high, with ICC values between 0.91 and 0.98. Changes seen in thermal scans when properly done are most likely due to actual physiological changes rather than equipment error.

Adult↗

AFLP typing of Staphylococcus epidermidis in multiple sequential blood cultures.

AFLP is a novel high-resolution PCR-based DNA fingerprinting method generating complex banding patterns that can be used for comparative analysis. In the present study, the applicability of AFLP in fingerprinting Staphylococcus epidermidis isolates was investigated. The criteria considered were stability of patterns, reproducibility, discriminatory capacity and consistency with epidemiological context. Repeated testing of strains and investigation of subcultures showed that AFLP patterns were reproducible and stable with an intrastrain similarity of S > or = 94% as determined by analysis of digitized patterns. Fifteen unrelated strains were heterogeneous, with a level ranging from 78-93%. The applicability of AFLP in epidemiological studies of S. epidermidis was tested on 11 sets of four blood isolates each, from 11 patients with suspected septicaemia. Nine sets had indistinguishable or highly similar AFLP patterns for each isolate per set, while two sets had heterogeneous patterns. These results show that AFLP has high discriminatory power for strain identification in S. epidermidis.

Bacteremia↗

The reproducibility of organ position using active breathing control (ABC) during liver radiotherapy.

PURPOSE: To evaluate the intrafraction and interfraction reproducibility of liver immobilization using active breathing control (ABC). METHODS AND MATERIALS: Patients with unresectable intrahepatic tumors who could comfortably hold their breath for at least 20 s were treated with focal liver radiation using ABC for liver immobilization. Fluoroscopy was used to measure any potential motion during ABC breath holds. Preceding each radiotherapy fraction, with the patient setup in the nominal treatment position using ABC, orthogonal radiographs were taken using room-mounted diagnostic X-ray tubes and a digital imager. The radiographs were compared to reference images using a 2D alignment tool. The treatment table was moved to produce acceptable setup, and repeat orthogonal verification images were obtained. The positions of the diaphragm and the liver (assessed by localization of implanted radiopaque intra-arterial microcoils) relative to the skeleton were subsequently analyzed. The intrafraction reproducibility (from repeat radiographs obtained within the time period of one fraction before treatment) and interfraction reproducibility (from comparisons of the first radiograph for each treatment with a reference radiograph) of the diaphragm and the hepatic microcoil positions relative to the skeleton with repeat breath holds using ABC were then measured. Caudal-cranial (CC), anterior-posterior (AP), and medial-lateral (ML) reproducibility of the hepatic microcoils relative to the skeleton were also determined from three-dimensional alignment of repeat CT scans obtained in the treatment position. RESULTS: A total of 262 fractions of radiation were delivered using ABC breath holds in 8 patients. No motion of the diaphragm or hepatic microcoils was observed on fluoroscopy during ABC breath holds. From analyses of 158 sets of positioning radiographs, the average intrafraction CC reproducibility (sigma) of the diaphragm and hepatic microcoil position relative to the skeleton using ABC repeat breath holds was 2.5 mm (range 1.8-3.7 mm) and 2.3 mm (range 1.2-3.7 mm) respectively. However, based on 262 sets of positioning radiographs, the average interfraction CC reproducibility (sigma) of the diaphragm and hepatic microcoils was 4.4 mm (range 3.0-6.1 mm) and 4.3 mm (range 3.1-5.7 mm), indicating a change of diaphragm and microcoil position relative to the skeleton over the course of treatment with repeat breath holds at the same phase of the respiratory cycle. The average population absolute intrafraction CC offset in diaphragm and microcoil position relative to skeleton was 2.4 mm and 2.1 mm respectively; the average absolute interfraction CC offset was 5.2 mm. Analyses of repeat CT scans demonstrated that the average intrafraction excursion of the hepatic microcoils relative to the skeleton in the CC, AP, and ML directions was 1.9 mm, 0.6 mm, and 0.6 mm respectively and the average interfraction CC, AP, and ML excursion of the hepatic microcoils was 6.6 mm, 3.2 mm, and 3.3 mm respectively. CONCLUSION: Radiotherapy using ABC for patients with intrahepatic cancer is feasible, with good intrafraction reproducibility of liver position using ABC. However, the interfraction reproducibility of organ position with ABC suggests the need for daily on-line imaging and repositioning if treatment margins smaller than those required for free breathing are a goal.

Humans↗

Measurement of tendon reflexes by surface electromyography in normal subjects.

A simple method for measuring the tendon reflexes was developed. A manually operated, electronic reflex hammer was applied that enabled measurement of the strength of tendon taps. Reflex responses were recorded by surface electromyography. Stimulus-response relations and latencies of tendon reflexes in the biceps, triceps, quadriceps and triceps surae were examined in 40 healthy subjects. A characteristic relation between stimulus strength and response amplitude was found which could be described by an empirical function. Latencies of both arm and leg reflexes were linearly related to the height of the subjects. Variations of reflex amplitudes within and between subjects were comparable with previous results obtained with more complicated techniques. Although repeatability of measurement of the amplitude is limited by the variability of reflexes, significant agreement was found between repeated measurements. Most reflex amplitudes were diminished during repeated examination after a short interval. Both measurement and clinical examination showed the frequent occurrence of left-right asymmetry of reflex amplitudes. These left-right differences were reproducible to a significant degree on repeated measurements after more than 2 years.

Adult↗

Development of stereotactically guided laser interstitial thermotherapy of breast cancer: in situ measurement and analysis of the temperature field in ex vivo and in vivo adipose tissue.

BACKGROUND AND OBJECTIVE: The size (0.5-1.0 cm) of early nonpalpable breast tumors currently detected by mammography and confirmed by stereotactic core biopsy is of the order of the penetration depth of near infrared photons in breast tissue. In principle, stereotactically biopsied tumors, therefore, could be safely and efficiently treated with laser thermotherapy. The aim of the current study is to confirm the controlled heating produced by clinically relevant power levels delivered with an interstitial laser fiber optic probe adapted for use with stereotactic mammography and biopsy procedures. STUDY DESIGN/MATERIALS AND METHODS: Temperature increases and the resultant thermal field produced by the irradiation of ex vivo (porcine and human) and in vivo (porcine) tissue models appropriate to the treatment of human breast tissue by using cw Nd:YAG laser radiation delivered with a interstitial fiber optic probe with a quartz diffusing tip, were recorded with an array of fifteen 23-gauge needle thermocouple probes connected to a laboratory computer-based data acquisition system. RESULTS: By using a stepwise decreasing power cycle to avoid tissue charring, acceptably symmetric thermal fields of repeatable volumetric dimensions were obtained. Reproducible thermal gradients and predictable tissue necrosis without carbonization could be induced in a 3-cm-diameter region around the fiber probe during a single treatment lasting only 3 minutes. The time-dependences of the temperature rise of the thermocouples surrounding the LITT probe were quantitatively modeled with simple linear functions during the applied laser heating cycles. CONCLUSION: Analysis of our experimental results show that reproducible, symmetric and predictable volumetric temperature increases in time can be reliably produced by interstitial laser thermotherapy.

Adipose Tissue↗

An in vitro validation of a commercially available metabolic cart using pediatric ventilator volumes.

Our objective was to determine the validity of the SensorMedics 2900 Metabolic Measurement Cart (SensorMedics Corp., Yorba Linda, CA) for use at pediatric ventilator volumes by designing an in vitro validation study. The study was performed using a previously described nitrogen dilution, carbon dioxide infusion technique in which known amounts of nitrogen (N2) and carbon dioxide (CO2) are infused while a test lung is ventilated. Metabolic measurements were made at tidal volumes (Vt) varying from 50-500 mL at a peak end-expiratory pressure (PEEP) of 0 and 12 cm H2O and at inspiratory oxygen fractions (Fi,O2s) ranging from 0.21-0.60. Each study was repeated six times to assess reproducibility of measurements. At a PEEP of 0 cm H2O, the measurement of oxygen consumption (V'O2), carbon dioxide production (V'CO2), and minute ventilation (V'E) were all within +/-10% of predicted values at all Fi,O2S except when Vt(t) was 50 mL. When PEEP was increased to 12 cm H2O, error in measurement of V'O2 increased and became unacceptably large as Fi,O2 was increased. Each study consisted of six trials, and results showed acceptable reproducibility as demonstrated by narrow 95% confidence intervals. For V(t)s of 100-500 mL, the metabolic monitor provided accurate and reproducible results under in vitro testing conditions when PEEP was low. Accurate results could not be obtained with a Vt of 50 mL or at elevated PEEP.

Calorimetry, Indirect↗

Pharmacokinetics of furosemide after three different single oral doses.

Furosemide solution was orally administered to 21 healthy adult males to determine dose proportionality over the dose range used and the reproducibility of disposition following a repeated dose. Furosemide solution was given in doses of 20, 40, and 80 mg, with the 40 mg dose repeated once. Blood was collected for 12 hours post-dose and urine for 24 hours. The maximum plasma concentrations resulting from 20, 40, and 80 mg doses were significantly different (p less than 0.05). Dose normalized maximum concentrations for the 20 and 80 mg doses were significantly different (p less than 0.05). Mean time to Cpmax was 50 minutes, with no differences observed among doses. Plasma AUCs were significantly different (p less than 0.05) for 20, 40, and 80 mg. Dose normalized AUCs were not significantly different. Mean amounts of furosemide in urine (Xu) were 9.62, 16.7, and 32.0 mg for the 20, 40, and 80 mg doses, respectively. These amounts were significantly different (p less than 0.05); dose normalized amounts were not significantly different. Renal clearances of furosemide following the three doses were not significantly different. Regressions of Cpmax, AUC and Xu on dose were significant. There were no significant differences in Cpmax, tmax, AUC or Xu for 40 mg given on two separate days. Renal clearance of furosemide was statistically different for 40 mg given on two separate days, but the difference was not clinically significant. The pharmacokinetics of furosemide are linear over the dosage range studied. Furosemide 40 mg given on two separate days results in similar disposition parameters.

Administration, Oral↗

Performance characteristics of an analytical procedure for determining chloramphenicol residues in muscle tissue by gas chromatography-electron capture detection.

Validation of an analytical method for determining chloramphenicol residues in muscle tissue by gas chromatography-electron capture detection (GC-ECD) was performed according to the latest European Union criteria for the analysis of veterinary drugs in food, laid down by Commission Decision 2002/657/EC. The method using the meta isomer of chloramphenicol as an internal standard proved to be very selective, specific to other related phenicols and accurate to within +3.6% at a concentration level of 8.9 microg/kg, as present in the certified reference material available. The correlation coefficient of the calibration curve was 0.9991. At all three fortification levels studied (0.3, 0.45 and 0.6 microg/kg), repeatability and intra-laboratory reproducibility were <8 and < or =9%, respectively. The decision limit (CCalpha) and detection capability (CCbeta) were 0.07 and 0.12 microg/kg, respectively. The validation results and the results of participation in an international inter-laboratory proficiency test indicate that the method presented is completely suited for regulatory control to screen and quantify chloramphenicol residues in various muscle tissues on a routine basis.

Animals↗

Determination of B2 and B6 vitamers in serum by capillary electrophoresis-molecular fluorescence-charge coupled detector.

A method for the separation by capillary zone electrophoresis and determination of vitamers of two important water-soluble vitamins, i.e., B2 and B6, is proposed here. The working conditions for optimal separation were obtained by a multivariate methodology in order to succeed in the best resolution in the shortest analysis time. The optimization of the buffer composition together with other variables, such as analysis and injection voltage, temperature of the capillary cassette, and injection time, resulted in a solution of 30 mM KH2PO4 adjusted to pH 8.5 with formic acid. Concerning the detection step, the target analytes were quantified by molecular fluorescence, for which two different detectors, a photomultiplier tube (PMT) and a charge-coupled detector (CCD), were compared in terms of resolution, sensitivity, and precision. A Xe-Hg lamp was used as an irradiation source in the two cases. The best option was the CCD, which provides three-dimensional electropherograms and enables to solve the overlapped peaks. Besides, the sensitivity of the CCD was similar to that of the PMT, due to the treatment data, obtaining limits of detection and quantification from 1.16 to 27.1 ng/g and from 3.83 to 89.4 ng/g, respectively. The method was applied to the serum samples for which a prior liquid-liquid extraction using ethanol in an acid medium was mandatory for eliminating the interferences and concentrating the analytes by a factor of 5. The rapidity of the analysis (13 min for the electrophoretic separation) and the excellent precision (repeatability and within-laboratory reproducibility between 2.86 and 4.11% and 7.03 and 8.45%, respectively, both expressed as relative standard deviation) demonstrated the capability of the proposed method for a clinical routine analysis.

Electrophoresis, Capillary↗