A Dilution Method to Mitigate Biotin Interference in Cardiac Troponin T Testing.

BACKGROUND: Oral biotin supplementation is known to interfere with biotin-streptavidin-based immunoassays, including Roche's fifth-generation cardiac troponin T (cTnT) assay, which plays a critical role in the diagnosis of myocardial infarction (MI). The utility of dilution, a quick and easy method to detect and remove interferences, has not been published for biotin interference. METHODS: Concentrations of cTnT were measured in pooled serum from clinical samples. Serum samples were supplemented with biotin to known concentrations, then cTnT concentrations were remeasured to assess for biotin interference. Samples were then diluted to assess for effective removal of biotin interference. RESULTS: At cTnT values near the critical reporting range for our institution (100 ng/L) we observed significant interference in measured values with added biotin concentrations above 50 ng/mL. In specimens without added biotin, autodilution at a 1:10 ratio yielded a mean 157% capture of measured cTnT, precluding the use of autodilution for detecting and mitigating biotin interference. A 1:10 dilution with serum containing 20-30 ng/L cTnT yielded a mean capture of 107%, which was suitable for detecting underlying biotin interference in supplemented samples. CONCLUSIONS: Biotin interference, at supraphysiologic concentrations, may create an artifactual reduction in measured cTnT to levels that could lead to delayed detection of an MI. Dilution with serum of known cTnT concentration of 20-30 ng/L is a fast and effective method to mitigate the analytical consequences of biotin interference.

Performance of Ceftolozane-Tazobactam Etest, MIC Test Strips, and Disk Diffusion Compared to Reference Broth Microdilution for ß-Lactam-Resistant Pseudomonas aeruginosa Isolates.

The performance characteristics of the ceftolozane-tazobactam (C-T) Etest (bioMérieux, Marcy l'Etoile, France), MIC test strips (MTS; Liofilchem, Italy), and disk diffusion (Hardy, Santa Ana, CA) were evaluated for a collection of 308 beta-lactam-resistant isolates of Pseudomonas aeruginosa recovered from three institutions in Los Angeles, CA. Reference testing was performed by the reference broth microdilution (rBMD) method. MIC and disk results were interpreted using Clinical and Laboratory Standards Institute breakpoints. Overall, 72.5% of the isolates were susceptible to C-T by rBMD. Etest and disk diffusion demonstrated acceptable performance, whereas MTS yielded a greater than acceptable percentage of minor errors. Categorical agreement was 96.8% for Etest, 87.0% for MTS, and 92.9% for disk diffusion. No very major errors were observed by any test, and no major errors (ME) were observed by Etest or disk diffusion. Two ME (0.9% of susceptible isolates) were observed by MTS. The incidence of minor errors was 3.2%, 12.3%, and 7.1% for Etest, MTS, and disk diffusion, respectively. Essential agreement (EA) for Etest was excellent, at 97.7%, whereas the MICs obtained by MTS tended to be 1 to 2 dilutions higher than those obtained by rBMD, with an EA of 87.0%.

Determination of serum calcium levels by 42Ca isotope dilution inductively coupled plasma mass spectrometry.

BACKGROUND: Serum calcium level is an important clinical index that reflects pathophysiological states. However, detection accuracy in laboratory tests is not ideal; as such, a high accuracy method is needed. METHODS: We developed a reference method for measuring serum calcium levels by isotope dilution inductively coupled plasma mass spectrometry (ID ICP-MS), using 42Ca as the enriched isotope. Serum was digested with 69% ultrapure nitric acid and diluted to a suitable concentration. The 44Ca/42Ca ratio was detected in H2 mode; spike concentration was calibrated by reverse IDMS using standard reference material (SRM) 3109a, and sample concentration was measured by a bracketing procedure. We compared the performance of ID ICP-MS with those of three other reference methods in China using the same serum and aqueous samples. RESULTS: The relative expanded uncertainty of the sample concentration was 0.414% (k=2). The range of repeatability (within-run imprecision), intermediate imprecision (between-run imprecision), and intra-laboratory imprecision were 0.12%-0.19%, 0.07%-0.09%, and 0.16%-0.17%, respectively, for two of the serum samples. SRM909bI, SRM909bII, SRM909c, and GBW09152 were found to be within the certified value interval, with mean relative bias values of 0.29%, -0.02%, 0.10%, and -0.19%, respectively. The range of recovery was 99.87%-100.37%. Results obtained by ID ICP-MS showed a better accuracy than and were highly correlated with those of other reference methods. CONCLUSIONS: ID ICP-MS is a simple and accurate candidate reference method for serum calcium measurement and can be used to establish and improve serum calcium reference system in China.

Harmonisation of serum dihydrotestosterone analysis: establishment of an external quality assurance program.

BACKGROUND: Serum dihydrotestosterone (DHT) is an important analyte for the clinical assessment of disorders of sex development. It is also reportedly a difficult analyte to measure. Currently, there are significant gaps in the standardisation of this analyte, including no external quality assurance (EQA) program available worldwide to allow for peer review performance of DHT. We therefore proposed to establish a pilot EQA program for serum DHT. METHODS: DHT was assessed in the 2015 Royal College of Pathologists of Australasia Quality Assurance Programs' Endocrine program material. The material's target (i.e. "true") values were established using a measurement procedure based on isotope dilution gas chromatography (GC) tandem mass spectrometry (MS/MS). DHT calibrator values were based on weighed values of pure DHT material (>97.5% purity) from Sigma. The allowable limits of performance (ALP) were established as ±0.1 up to 0.5 nmol/L and ±15% for targets >0.5 nmol/L. RESULTS: Target values for the six levels of RCPAQAP material for DHT ranged from 0.02 to 0.43 nmol/L (0.01-0.12 ng/mL). The material demonstrated linearity across the six levels. There were seven participating laboratories for this pilot study. Results of the liquid chromatography (LC) MS/MS methods were within the ALP; whereas the results from the immunoassay methods were consistently higher than the target values and outside the ALP. CONCLUSIONS: This report provides the first peer comparison of serum DHT measured by mass spectrometry (MS) and immunoassay laboratories. Establishment of this program provides one of the pillars to achieve method harmonisation. This supports accurate clinical decisions where DHT measurement is required.

Evaluation of an Isotope Dilution HPLC Tandem Mass Spectrometry Candidate Reference Measurement Procedure for Total 17-ß Estradiol in Human Serum.

The inaccuracy of 17-ß estradiol (E2) measurements affects its use as a biomarker in patient care and research. Clinical and research communities called for accurate and standardized E2 measurements. Reference Measurement Procedures (RMPs), part of the CDC Hormone Standardization Program (HoSt), are essential in addressing this need and ensuring that methods are accurate and comparable across testing systems, laboratories, and over time. A candidate RMP (cRMP) was developed for the measurement of total E2 in serum using liquid chromatography-tandem mass spectrometry (LC-MS/MS) without derivatization. The cRMP meets suggested performance criteria for accuracy and precision through the use of isotope dilution, calibrator bracketing, and gravimetric measurements. The cRMP demonstrated high agreement with certified reference materials (no significant bias to BCR576, 577, and 578) and established RMPs (slope 1.00, 95% CI 1.00-1.01; intercept 0.02, 95% CI -0.01 to 0.06). The cRMP is highly precise with intra-assay, interassay, and total percent CVs of 2.7%, 1.3%, and 2.4%, respectively. A higher specificity was achieved by measuring E2 without derivatization, compared to methods using derivatization agents. The cRMP can serve as a higher-order standard for establishing measurement traceability and provides an accuracy base against which routine methods can be compared in HoSt.

Replications of fundamental research models in ultra high dilutions 1994 and 2015--update on a bibliometric study.

INTRODUCTION: This paper focuses exclusively on experimental models with ultra high dilutions (i.e. beyond 10(-23)) that have been submitted to replication scrutiny. It updates previous surveys, considers suggestions made by the research community and compares the state of replication in 1994 with that in 2015. METHODS: Following literature research, biochemical, immunological, botanical, cell biological and zoological studies on ultra high dilutions (potencies) were included. Reports were grouped into initial studies, laboratory-internal, multicentre and external replications. Repetition could yield either comparable, or zero, or opposite results. The null-hypothesis was that test and control groups would not be distinguishable (zero effect). RESULTS: A total of 126 studies were found. From these, 28 were initial studies. When all 98 replicative studies were considered, 70.4% (i.e. 69) reported a result comparable to that of the initial study, 20.4% (20) zero effect and 9.2% (9) an opposite result. Both for the studies until 1994 and the studies 1995-2015 the null-hypothesis (dominance of zero results) should be rejected. Furthermore, the odds of finding a comparable result are generally higher than of finding an opposite result. Although this is true for all three types of replication studies, the fraction of comparable studies diminishes from laboratory-internal (total 82.9%) to multicentre (total 75%) to external (total 48.3%), while the fraction of opposite results was 4.9%, 10.7% and 13.8%. Furthermore, it became obvious that the probability of an external replication producing comparable results is bigger for models that had already been further scrutinized by the initial researchers. CONCLUSIONS: We found 28 experimental models which underwent replication. In total, 24 models were replicated with comparable results, 12 models with zero effect, and 6 models with opposite results. Five models were externally reproduced with comparable results. We encourage further replications of studies in order to learn more about the model systems used.

Validation of Bioelectrical Impedance Spectroscopy to Measure Total Body Water in Resistance-Trained Males.

The three-compartment (3-C) model of physique assessment (fat mass, fat-free mass, water) incorporates total body water (TBW) whereas the two-compartment model (2-C) assumes a TBW of 73.72%. Deuterium dilution (D2O) is the reference method for measuring TBW but is expensive and time consuming. Multifrequency bioelectrical impedance spectroscopy (BIS SFB7) estimates TBW instantaneously and claims high precision. Our aim was to compare SFB7 with D2O for estimating TBW in resistance trained males (BMI >25kg/m2). We included TBWBIS estimates in a 3-C model and contrasted this and the 2-C model against the reference 3-C model using TBWD2O. TBW of 29 males (32.4 ± 8.5 years; 183.4 ± 7.2 cm; 92.5 ± 9.9 kg; 27.5 ± 2.6 kg/m2) was measured using SFB7 and D2O. Body density was measured by BODPOD, with body composition calculated using the Siri equation. TBWBIS values were consistent with TBWD2O (SEE = 2.65L; TE = 2.6L) as were %BF values from the 3-C model (BODPOD + TBWBIS) with the 3-C reference model (SEE = 2.20%; TE = 2.20%). For subjects with TBW more than 1% from the assumed 73.72% (n = 16), %BF from the 2-C model differed significantly from the reference 3-C model (Slope 0.6888; Intercept 5.093). The BIS SFB7 measured TBW accurately compared with D2O. The 2C model with an assumed TBW of 73.72% introduces error in the estimation of body composition. We recommend TBW should be measured, either via the traditional D2O method or when resources are limited, with BIS, so that body composition estimates are enhanced. The BIS can be accurately used in 3C equations to better predict TBW and BF% in resistance trained males compared with a 2C model.

Simultaneous determination of 11 ß-agonists in human urine using high-performance liquid chromatography/tandem mass spectrometry with isotope dilution.

The misuse of ß-agonists constitutes a potential risk to public health and has been forbidden in many countries. In this study, we describe a method for specific, sensitive and rapid detection of ß-agonists in human urine. Urine samples were extracted with ethyl acetate, without any additional purification step, and analyzed by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS-MS) with Clenbuterol-D9 and Salbuterol-D3 as internal standards. The intra- and interday precision values of the method were all <5.60% and the accuracy ranged from 94.5 to 109%. Extraction recovery for 11 ß-agonists varied from 66.7 to 108%. One UPLC-MS-MS analysis could be completed within 12 min and the limits of detection for 11 ß-agonists were 0.1 ng/mL in the experiment. ß-Agonists in human urines from 24 volunteers were analyzed by our validated method and 1.70 ng/mL salbutamol was detected in one volunteer. The application of UPLC-MS-MS method in ß-agonists detection of human urine will be helpful in veterinary control of ß-agonists and for studying the effect of ß-agonists on human health.

Measurement procedure for glucose in serum using liquid chromatography isotope dilution--tandem mass spectrometry (LC-ID-MS/MS).

BACKGROUND: We developed and validated a measurement procedure for glucose using liquid chromatography-isotope dilution tandem mass spectrometry. The proposed method is intended to be used for setting target values in EQAS samples and for certification of glucose reference materials, including those in biological matrices. METHODS: Serum samples were spiked with internal standard 13C6 D-glucose. Protein precipitation was performed with ethanol. The samples were vortexed and centrifuged. An aliquot of the supernatant was evaporated to dryness, the residue dissolved in elution buffer and injected into the LC-MS/MS system. Measurements were performed in the positive ion mode monitoring the Cs+ adducts for glucose at m/z 313 --> 132.9 and m/z 319 --> 132.9 for the internal standard. RESULTS: The coefficient of variation (CV) of the measurement procedure for lyophilized, liquid, and fresh serum samples was between 0.27 and 1.77%. The bias from certified target values for NIST reference materials was < or = 0.62%. A Deming regression comparison demonstrated a good correlation of results obtained with the proposed LC-ID-MS/MS method and target values obtained in the internationally accepted IFCC-RELA ring trial using JCTLM-recognized reference measurement procedures. CONCLUSIONS: The proposed LC-ID-MS/MS measurement procedure with traceability to SI units shows excellent accuracy and precision and is suitable for use as reference measurement procedure for certification of target values in lyophilized and fresh serum samples.

Biosynthesis of 15N-labeled cylindrospermopsin and its application as internal standard in stable isotope dilution analysis.

Cylindrospermopsin (CYN) is a cyanobacterial toxin associated with human and animal poisonings. Due to its toxicity in combination with its widespread occurrence, the development of reliable methods for selective, sensitive detection and accurate quantification is mandatory. Liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis using stable isotope dilution analysis (SIDA) represents an ideal tool for this purpose. U-[(15)N5]-CYN was synthesized by culturing Aphanizomenon flos-aquae in Na(15)NO3-containing cyanobacteria growth medium followed by a cleanup using graphitized carbon black columns and mass spectrometric characterization. Subsequently, a SIDA-LC-MS/MS method for the quantification of CYN in freshwater and Brassica matrices was developed showing satisfactory performance data. The recovery ranged between 98 and 103 %; the limit of quantification was 15 ng/L in freshwater and 50 µg/kg dry weight in Brassica samples. The novel SIDA was applied for CYN determination in real freshwater samples as well as in kale and in vegetable mustard exposed to toxin-containing irrigation water. Two of the freshwater samples taken from German lakes were found to be CYN-contaminated above limit of quantification (17.9 and 60.8 ng/L). CYN is systemically available to the examined vegetable species after exposure of the rootstock leading to CYN mass fractions in kale and vegetable mustard leaves of 15.0 µg/kg fresh weight and 23.9 µg/kg fresh weight, respectively. CYN measurements in both matrices are exemplary for the versatile applicability of the developed method in environmental analysis.

Application of the reference method isotope dilution gas chromatography mass spectrometry (ID/GC/MS) to establish metrological traceability for calibration and control of blood glucose test systems.

Self-monitoring of blood glucose (BG) by means of handheld BG systems is a cornerstone in diabetes therapy. The aim of this article is to describe a procedure with proven traceability for calibration and evaluation of BG systems to guarantee reliable BG measurements. Isotope dilution gas chromatography mass spectrometry (ID/GC/MS) is a method that fulfills all requirements to be used in a higher-order reference measurement procedure. However, this method is not applicable for routine measurements because of the time-consuming sample preparation. A hexokinase method with perchloric acid (PCA) sample pretreatment is used in a measurement procedure for such purposes. This method is directly linked to the ID/GC/MS method by calibration with a glucose solution that has an ID/GC/MS-determined target value. BG systems are calibrated with whole blood samples. The glucose levels in such samples are analyzed by this ID/GC/MS-linked hexokinase method to establish traceability to higher-order reference material. For method comparison, the glucose concentrations in 577 whole blood samples were measured using the PCA-hexokinase method and the ID/GC/MS method; this resulted in a mean deviation of 0.1%. The mean deviation between BG levels measured in >500 valid whole blood samples with BG systems and the ID/GC/MS was 1.1%. BG systems allow a reliable glucose measurement if a true reference measurement procedure, with a noninterrupted traceability chain using ID/GC/MS linked hexokinase method for calibration of BG systems, is implemented. Systems should be calibrated by means of a traceable and defined measurement procedure to avoid bias.

A regional survey of serum creatinine measurement methods and estimated glomerular filtration rate reporting.

BACKGROUND: Only limited data are available on the diffusion of isotope dilution mass spectrometry (IDMS)-traceable methods used for serum creatinine measurement and on estimated glomerular filtration rate (eGFR) reporting. METHODS: A questionnaire was addressed to accredited laboratories in Lombardy, Italy, including the following issues: method of creatinine measurement, instrument model, IDMS calibration traceability, reference intervals reported by sex and age, eGFR reporting, eGFR formula used and information about the eGFR value reported in patient records. A parallel questionnaire was addressed to nephrology centers and included the following: knowledge of methods for serum creatinine measurement in their center, usefulness of eGRF reporting and opinions on the need for educational initiatives. RESULTS: Seventy-two percent of 72 laboratories and 89% of 47 nephrology centers responded to the questionnaires. Among the methods used for serum creatinine measurement, 87% were IDMS traceable and 30% were enzymatic. Reference intervals were differentiated by sex and by age in 90% and 42%, respectively. Laboratories reported eGFR in 35% and only when requested in 13%. eGFR was calculated by the Modification of Diet in Renal Disease (MDRD) Study and Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) equations in 88% and 12% of laboratories, respectively, and reporting was accompanied by information on the interpretation of values in 62%. Among nephrologists, 64% thought eGFR reporting useful, 29% were concerned with an excess of unnecessary requests for consultations and 95% expressed a favorable opinion of educational initiatives. CONCLUSION: Our survey highlights the need for further improvement in serum creatinine measurement and reporting, and for coordinated interventions involving all major stakeholders.

Bioelectrical impedance with different equations versus deuterium oxide dilution method for the inference of body composition in healthy older persons.

BACKGROUND: There is no consensus regarding the accuracy of bioimpedance for the determination of body composition in older persons. OBJECTIVE: This study aimed to compare the assessment of lean body mass of healthy older volunteers obtained by the deuterium dilution method (reference) with those obtained by two frequently used bioelectrical impedance formulas and one formula specifically developed for a Latin-American population. DESIGN: A cross-sectional study. PARTICIPANTS: Twenty one volunteers were studied, 12 women, with mean age 72±6.7 years. SETTING: Urban community, Ribeirão Preto, Brazil. MEASUREMENT: Fat free mass was determined, simultaneously, by the deuterium dilution method and bioelectrical impedance; results were compared. In bioelectrical impedance, body composition was calculated by the formulas of Deuremberg, Lukaski and Bolonchuck and Valencia et al. RESULTS: Lean body mass of the studied volunteers, as determined by bioelectrical impedance was 37.8±9.2 kg by the application of the Lukaski e Bolonchuk formula, 37.4±9.3 kg (Deuremberg) and 43.2±8.9 kg (Valencia et. al.). The results were significantly correlated to those obtained by the deuterium dilution method (41.6±9.3 Kg), with r=0.963, 0.932 and 0.971, respectively. Lean body mass obtained by the Valencia formula was the most accurate. CONCLUSION: In this study, lean body mass of older persons obtained by the bioelectrical impedance method showed good correlation with the values obtained by the deuterium dilution method. The formula of Valencia et al., developed for a Latin-American population, showed the best accuracy.

Performance of the AOAC use-dilution method with targeted modifications: collaborative study.

The U.S. Environmental Protection Agency (EPA), in collaboration with an industry work group, spearheaded a collaborative study designed to further enhance the AOAC use-dilution method (UDM). Based on feedback from laboratories that routinely conduct the UDM, improvements to the test culture preparation steps were prioritized. A set of modifications, largely based on culturing the test microbes on agar as specified in the AOAC hard surface carrier test method, were evaluated in a five-laboratory trial. The modifications targeted the preparation of the Pseudomonas aeruginosa test culture due to the difficulty in separating the pellicle from the broth in the current UDM. The proposed modifications (i.e., the modified UDM) were compared to the current UDM methodology for P. aeruginosa and Staphylococcus aureus. Salmonella choleraesuis was not included in the study. The goal was to determine if the modifications reduced method variability. Three efficacy response variables were statistically analyzed: the number of positive carriers, the log reduction, and the pass/fail outcome. The scope of the collaborative study was limited to testing one liquid disinfectant (an EPA-registered quaternary ammonium product) at two levels of presumed product efficacies, high and low. Test conditions included use of 400 ppm hard water as the product diluent and a 5% organic soil load (horse serum) added to the inoculum. Unfortunately, the study failed to support the adoption of the major modification (use of an agar-based approach to grow the test cultures) based on an analysis of method's variability. The repeatability and reproducibility standard deviations for the modified method were equal to or greater than those for the current method across the various test variables. However, the authors propose retaining the frozen stock preparation step of the modified method, and based on the statistical equivalency of the control log densities, support its adoption as a procedural change to the current UDM. The current UDM displayed acceptable responsiveness to changes in product efficacy; acceptable repeatability across multiple tests in each laboratory for the control counts and log reductions; and acceptable reproducibility across multiple laboratories for the control log density values and log reductions. Although the data do not support the adoption of all modifications, the UDM collaborative study data are valuable for assessing sources of method variability and a reassessment of the performance standard for the UDM.

Development and validation of a reference measurement procedure for certification of phenytoin, phenobarbital, lamotrigine, and topiramate in human serum using isotope-dilution liquid chromatography/tandem mass spectrometry.

Phenytoin (PHT), phenobarbital (PHB), lamotrigine (LTG), and topiramate (TPM) are some of the most widely used antiepileptic drugs (AEDs). Monitoring of their concentrations in serum is important for the treatment of epilepsy. A reference measurement procedure (RMP) for certification of PHT, PHB, LTG, and TPM in serum has been developed and critically evaluated. Isotopically labeled compounds of PHT, PHB, LTG, and TPM are used as internal standards for the four AEDs. The four drugs and their respective labeled internal standards are simultaneously extracted from serum using solid-phase extraction prior to reversed-phase liquid chromatography-tandem mass spectrometry (LC-MS/MS). Chromatographic separation was performed using a C(18) column. Electrospray ionization (ESI) in the positive ion mode for PHT and LTG, and in the negative ion mode for PHB and TPM were used. The recovery of AEDs added to serum (accuracy of the extraction method) was evaluated by recovery studies of measuring the four drugs in spiked samples with known drug levels. The recoveries of the added drugs ranged from 98.6% to 102.0%. The absolute recoveries (extraction efficiencies) of the four drugs with this method ranged from 97% to 100%. Excellent repeatability was obtained for the four drugs with between-set coefficients of variation (CVs) within 1%. The type B components estimates are conservatively large and are considerably larger than the type A component. Therefore, we use the usual metrological expansion factor of 2 to provide an approximate 95% coverage interval. The relative expanded uncertainties for the four AEDs ranged from 2.3% to 2.4%. This LC-MS/MS RMP for PHT, PHB, LTG, and TPM in serum demonstrating good accuracy and precision can be used to assess the accuracy of routine methods used in clinical laboratories.

Rapid speciation and quantification of selenium compounds by HPLC-ICP MS using multiple standards labelled with different isotopes.

Speciation analysis using high-performance liquid chromatography-inductively coupled plasma mass spectrometry (HPLC-ICP MS) is now commonly used to investigate metabolic and toxicological aspects of some metals and metalloids. We have developed a rapid method for simultaneous identification and quantification of metabolites of selenium (Se) compounds using multiple standards labelled with different isotopes. A mixture of the labelled standards was spiked in a selenised garlic extract and the sample was subjected to speciation analysis by HPLC-ICP MS. The selenised garlic contains γ-glutamyl-methylselenocysteine, methylselenocysteine, and selenomethionine and the concentrations of those Se compounds were 723.8, 414.8, and 310.7 ng Se ml(-1), respectively. The isotopically labelled standards were also applied to the speciation of Se in rat urine. Selenate, methylselenonic acid, selenosugar, and trimethyselenium ions were found to be excreted by the present speciation procedure. Multiple standards labelled with different stable isotopes enable high-throughput identification and quantitative measurements of Se metabolites.

An inline QC method for determining serial dilution performance of DMSO-based systems.

Serial dilution of compounds solubilized in dimethylsulfoxide (DMSO) for dose-response curves is a common method for efficacy analysis of potential drug candidates. In general, serial dilution methods are particularly prone to error propagation because each dilution is dependent on the previous concentration. Moreover, assumptions about quality control parameters (i.e., dye linearity) can lead to an erroneous process. Here, an inline performance measurement is sought to improve the precision and accuracy of dilution plates. Sulforhodamine 101 (S101) dye is introduced as the quantitative fluorometric method of choice for DMSO-based systems. Although S101 in DMSO behaves in a nonlinear fashion over its detectable range, we account for this with a direct calibration method that includes every point of the dilution template. This report contains dye selection rationale for the S101 dye and its use in quantifying the performance of 96- and 384-well dilution protocols as tested on five identical instruments.

Comparative in vitro antimicrobial activities of torezolid (TR-700), the active moiety of a new oxazolidinone, torezolid phosphate (TR-701), determination of tentative disk diffusion interpretive criteria, and quality control ranges.

This study assessed the spectrum of activity of torezolid (TR-700), the active moiety of torezolid phosphate (TR-701), and proposes tentative MIC and disk diffusion breakpoints as well as quality control ranges. The in vitro susceptibilities of 1,096 bacterial isolates, representing 23 different species or phenotypic groups, were determined for torezolid, linezolid, cefotaxime, and levofloxacin using Clinical and Laboratory Standards Institute (CLSI) broth microdilution MICs, minimum bactericidal concentrations (MBCs), agar dilution, and disk diffusion testing methods. Torezolid was very active against the majority of Gram-positive strains, including methicillin-susceptible and -resistant Staphylococcus aureus (MIC(50) = 0.25 microg/ml, MIC(90)

Applicability of common reference intervals for serum creatinine concentrations to the Croatian population.

BACKGROUND: In accordance with an ongoing activity for worldwide harmonization based on traceability in laboratory methods, the goal of this study was to validate the applicability of recommended "common" reference intervals for serum creatinine concentrations using a specific enzymatic method to the Croatian population. METHODS: The reference group consisted of 240 healthy subjects (120 males and 120 females), between 18 and 74 years of age (median 57 years), who were selected in accordance with the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) recommendations. Creatinine in serum was measured using the creatinine enzymatic assay (Olympus OSR61204) that was standardized to the isotopic dilution mass spectrometry (IDMS) method and National Institute of Standards and Technology (NIST) Standard Reference Material (SRM) 967. In addition, creatinine was measured using a kinetic Jaffe method (Olympus OSR6178) standardized to NIST SRM 909b level 2 standard. RESULTS: Method comparison between enzymatic creatinine (x) and the Jaffe kinetic method (y) gave the following P/B equation for the entire group (n=240): y=1.00x+17.00; r=0.968. Reference intervals for serum creatinine (central 95th percentiles) obtained using the enzymatic creatinine method ranged from 54 to 107 micromol/L for males and from 50 to 93 micromol/L for females. The IFCC recommended common reference intervals for global applications are 64-104 micromol/L and 49-90 micromol/L for males and females, respectively. CONCLUSIONS: Comparability of obtained results confirmed the applicability of recently recommended "common" reference intervals to the Croatian population for all laboratories measuring serum creatinine concentrations using enzymatic methods traceable to the IDMS method and NIST SRM 967.