Performance evaluation of a turbidimetric cystatin C assay on different high-throughput platforms.

OBJECTIVE: The goal with this study was to evaluate the analytical performance of a new cystatin C immunoassay (Tina-quant a Cystatin C, Roche Diagnostics GmbH). The evaluation was carried out at four centers according to a standardized protocol. MATERIAL AND METHODS: The Tina-quant a Cystatin C is a latex particle-enhanced immunoturbidimetric assay. Roche cobas 6000, MODULAR ANALYTICS SWA and COBAS INTEGRA instruments were included in the study. Method comparison studies were carried out against two turbidimetric methods (Dako Cystatin C, Gentian Cystatin C), and one nephelometric method (Siemens N-Latex Cystatin C). RESULTS: Linearity was proven throughout the measuring range from 0.4 to 8 mg/L. Within-run CVs ranged from 0.7-2.8%, and total CVs from 1.4-4.7 % (concentration range 0.6-3.9 mg/L). Comparable results were obtained with paired serum and Li-heparinate plasma samples. Good agreement was achieved in the comparisons between the Tina-quant a Cystatin C assay and the other commercially available cystatin C assays, two different turbidimetric methods (slope range 0.88-1.04, intercept < 0.17 mg/L, r > or = 0.993) and one nephelometric assay (slope range 0.90-1.05, intercept < 0.21 mg/L, r > or = 0.986). CONCLUSIONS: The Tina-quant a Cystatin C assay was shown to be precise and accurate with proven linearity over the measuring range. Good comparability was obtained with other commercially available assays for the determination of cystatin C. The Tina-quant a Cystatin C assay is very well suited for clinical use on routine clinical chemistry analysers to detect renal dysfunction with a 24 h availability.

Therapeutic drug monitoring and drugs of abuse testing on the cobas 6000 analyzer series: analytical performance under routine-like conditions.

BACKGROUND: The analytical performance of the clinical chemistry module c 501 (cobas 6000 analyzer series) was evaluated for therapeutic drug monitoring and drugs of abuse testing using a spectrum of representative assays. Particular attention was paid to potential interactions between reagents using a simulated routine workload. METHODS: Within-run and total imprecision were assessed using a selection of representative reagents. Deviation from a consensus mean was tested using samples from a proficiency testing scheme. Method comparison using routine samples was carried out against the MODULAR ANALYTICS SWA and COBAS INTEGRA 800 analysis systems. RESULTS: Total coefficients of variation (CV) ranged from 1.9% to 7.8% for individual drugs, and from 3.2% to 8.6% for drugs of abuse testing. Results from proficiency test samples were between 81% and 125% of the consensus mean for therapeutic drugs. Method comparisons (Passing-Bablok regression) showed overall good comparability to MODULAR ANALYTICS SWA and COBAS INTEGRA 800 systems, with slopes from 0.93 to 1.17 and correlation coefficients r > 0.98. Imprecision in a simulated routine run was tested using a total of 42 methods (10 therapeutic drug monitoring, 9 drugs of abuse testing, 3 enzymes, 12 substrates, 8 specific protein assays). Imprecision in the reference batch run ranged from 0.7% to 5.0% CV for therapeutic drug monitoring assays, except for digoxin (DIG) (7.3%), and from 0.9% to 7.7% for drugs of abuse testing. The CVs of general clinical chemistry and specific protein tests were within the expected limits of 2% and 4%. CV changes in the simulated routine run were within the expected limits for most assays. Negative DeltaCVs (> or = 2%) for DIG, digitoxin (DIGIT), cannabinoids (THC), and phencyclidine (PCP) may indicate improved performance when running these assays in a simulated routine operation. A positive DeltaCV (> or = 3%) was found for amphetamines (AMPHs). CONCLUSIONS: In conclusion, the cobas c 501 module seems to be well-suited for routine use as consolidated workstation. Except for a potential interaction with AMPH, as indicated by the positive DeltaCV, no significant interferences from different reagents could be observed during this study.

Clinical utility of a new enzymatic assay for determination of mycophenolic acid in comparison with an optimized LC-MS/MS method.

The goal of this study was to investigate the clinical utility of a new enzymatic assay for use on COBAS INTEGRA systems (Roche Total MPA assay). From 134 patients, plasma mycophenolic acid (MPA) concentrations were measured with both the enzymatic method and a validated liquid chromatography with tandem mass spectrometry (LC-MS/MS) procedure, to compare these assays. The test principle of the enzymatic assay is inhibition of inosine monophosphate dehydrogenase. Method comparison studies revealed good agreement of results (r > 0.99), overall and in patients with delayed graft function or hypoalbuminemia. MPA area under the concentration-time curve (AUCs) obtained with LC-MS/MS (x) and the enzymatic method (y) compared excellent in patients on cyclosporine (y = 1.04x - 1.05, r = 0.992) or tacrolimus (y = 1.02x - 0.63, r = 0.987). MPA exposure determined with either method at different time points after transplantation agreed well (eg, 25th/50th/75th percentile of day 10 AUCs-LC-MS/MS: 25.8/33.8/45.2 versus enzymatic assay: 26.2/34.4/45.3 mg.h/L). AUCs calculated for both methods were lower at the first 3 time points in patients on cyclosporine compared with tacrolimus (week 4 median cyclosporine/tacrolimus: LC-MS/MS 39.6/56.4 versus enzymatic assay 40.5/56.0 mg.h/L). Both LC-MS/MS and the enzymatic methods revealed a tendency toward lower AUCs and predose levels in patients with biopsy-proven acute rejection (BPAR) (day 10 median: 0.9 mg/L with BPAR and 1.7 mg/L without BPAR). The Roche Total MPA assay is a reliable alternative to LC-MS/MS. It can be applied in the clinical setting allowing for easy, fast, and optimized patient management.

Multicenter evaluation of a new inosine monophosphate dehydrogenase inhibition assay for quantification of total mycophenolic acid in plasma.

The performance characteristics of a new inosine monophosphate dehydrogenase inhibition assay for the quantification of total mycophenolic acid (MPA) in plasma (Roche Diagnostics) were assessed in a multicenter evaluation. Validation data were collected from four institutions. Within-run and total imprecision were acceptable (n = 21 for each of 7 materials, coefficients of variation ranging 0.7-9.6%). The lower limit of quantification was 0.31 mg/L. The assay was linear from 0.31 to 15.0 mg/L. Method comparison with validated high-performance liquid chromatography with ultraviolet light or liquid chromatography tandem mass spectrometry methods showed good agreement (coefficients of correlation 0.974-0.994, slopes 1.01-1.17, intercepts -0.17 to 0.06). There was no difference found between results from different transplant types (cardiac vs. renal) or comedications (cyclosporine vs. tacrolimus). The recovery of samples from a proficiency testing scheme was acceptable. The cross-reactivity of AcMPAG, an in vitro active metabolite of MPA, was examined by adding AcMPAG to a pool of patient samples and subsequent quantification. MPA overestimation by AcMPAG cross-reactivity was found to be low (<5%). Thus, this interference is expected to be clinically irrelevant. In conclusion, the Roche Total MPA assay is a promising alternative for MPA quantification where chromatographic methods are not available.

Increasing Efficiency and Quality by Consolidation of Clinical Chemistry and Immunochemistry Systems with MODULAR ANALYTICS SWA.

MODULAR ANALYTICS Serum Work Area (in USA Integrated MODULAR ANALYTICS, MODULAR ANALYTICS is a trademark of a member of the Roche Group) represents a further approach to automation in the laboratory medicine. This instrument combines previously introduced modular systems for the clinical chemistry and immunochemistry laboratory and allows customised combinations for various laboratory workloads. Functionality, practicability, and workflow behaviour of MODULAR ANALYTICS Serum Work Area were evaluated in an international multicenter study at six laboratories. Across all experiments, 236000 results from 32400 samples were generated using 93 methods. Simulated routine testing which included provocation incidents and anomalous situations demonstrated good performance and full functionality. Heterogeneous immunoassays, performed on the E-module with the electrochemiluminescence technology, showed reproducibility at the same level of the general chemistry tests, which was well within the clinical demands. Sample carryover cannot occur due to intelligent sample processing. Workflow experiments for the various module combinations, with menus of about 50 assays, yielded mean sample processing times of <38 minutes for combined clinical chemistry and immunochemistry requests; <50 minutes including automatically repeated samples. MODULAR ANALYTICS Serum Work Area offered simplified workflow by combining various laboratory segments. It increased efficiency while maintaining or even improving quality of laboratory processes.

AmpliChip CYP450 GeneChip: a new gene chip that allows rapid and accurate CYP2D6 genotyping.

Methods for Cytochrome P450-2D6 (CYP2D6) genotyping are often time-consuming and laborious, which can restrict their use in pretherapeutic screening programs. Gene chip technology could overcome this problem. The aim of this study was to evaluate CYP2D6 genotyping by a new improved gene chip compared to a PCR-RFLP method. AmpliChip CYP450 GeneChip(R) (AmpliChip) is a microarray hybridization method for genotyping CYP2D6 and CYP2C19. One hundred fifty-nine DNA samples were genotyped both by AmpliChip as well as by PCR-RFLP and, where applicable, by a SNaPshot technique which detects single nucleotide polymorphisms based on the single base extension principle. In 152 of the 159 samples, CYP2D6 genotypes determined with the AmpliChip were in accordance with the results of PCR-RFLP. All seven discrepant samples had gene duplications and were subjected to SNaPshot analysis. SNaPshot results concurred with those of the AmpliChip for six out of seven samples. In the one divergent result, DNA sequencing confirmed that the AmpliChip had assigned the correct genotype. In conclusion, AmpliChip is a highly reliable method for CYP2D6 genotyping that allows the correct determination of all relevant CYP2D6 alleles in one single run. It therefore represents a very efficient and fast method, offering new perspectives for the application of pharmacogenetics in clinical medicine.

Automated cerebrospinal fluid cytology: limitations and reasonable applications.

OBJECTIVE: To evaluate the precision and clinical applicability of the Bayer ADVIA 120 cytometer (Bayer Healthcare, Fernwald, Germany) for cerebrospinal fluid (CSF) cell count and differentiation. STUDY DESIGN: One hundred six analyses of CSF from 98 patients by the ADVIA 120 were compared with routine cell count and microscopic differentiation. Correlation coefficients were calculated. RESULTS: In general, the total cell counts of both methods correlated well. The best correlations were seen at higher cell counts, > or = 100 cells per microliter with < 100 erythrocytes per microliter. The best correlations of cell differentiation were seen for lymphocytes and neutrophils, while the results for monocytes and eosinophils were less precise. In some cases, considerable differences between automated and microscopic cell counts and differentiation were seen that were relevant to clinical decision making. The detection of pathologic cell types, such as hemosiderophages, mitoses and neoplastic cells, was not provided by automated cytometry. CONCLUSION: When experienced personnel are not available, a preliminary cell count and differentiation between neutrophilic and lymphocytic reactions by automated cytometry may be valuable in allowing initial therapeutic decision making. Since the detection of pathologic cell types is not provided and the precision at low cell counts is only moderate, a personal microscopic evaluation of each sample is still indispensable to avoid misdiagnoses.

MODULAR ANALYTICS: A New Approach to Automation in the Clinical Laboratory.

MODULAR ANALYTICS (Roche Diagnostics) (MODULAR ANALYTICS, Elecsys and Cobas Integra are trademarks of a member of the Roche Group) represents a new approach to automation for the clinical chemistry laboratory. It consists of a control unit, a core unit with a bidirectional multitrack rack transportation system, and three distinct kinds of analytical modules: an ISE module, a P800 module (44 photometric tests, throughput of up to 800 tests/h), and a D2400 module (16 photometric tests, throughput up to 2400 tests/h). MODULAR ANALYTICS allows customised configurations for various laboratory workloads. The performance and practicability of MODULAR ANALYTICS were evaluated in an international multicentre study at 16 sites. Studies included precision, accuracy, analytical range, carry-over, and workflow assessment. More than 700 000 results were obtained during the course of the study. Median between-day CVs were typically less than 3% for clinical chemistries and less than 6% for homogeneous immunoassays. Median recoveries for nearly all standardised reference materials were within 5% of assigned values. Method comparisons versus current existing routine instrumentation were clinically acceptable in all cases. During the workflow studies, the work from three to four single workstations was transferred to MODULAR ANALYTICS, which offered over 100 possible methods, with reduction in sample splitting, handling errors, and turnaround time. Typical sample processing time on MODULAR ANALYTICS was less than 30 minutes, an improvement from the current laboratory systems. By combining multiple analytic units in flexible ways, MODULAR ANALYTICS met diverse laboratory needs and offered improvement in workflow over current laboratory situations. It increased overall efficiency while maintaining (or improving) quality.

Multicentre performance evaluation of the E170 module for modular analytics.

The E170 module was evaluated at 13 sites in an international multicentre study. The objective of the study was to assess the analytical performance of 49 analytes, and to collect feedback on the system's reliability and practicability. The typical, within-run coefficients of variation (CVs) for most of the quantitative assays ranged between 1 and 2% while a range of 2-4% was achieved with the infectious disease methods. Total precision CVs were found to be within the manufacturer's expected performance ranges, demonstrating good concordance of the system's measuring channels and a high reproducibility during the 2-4-week trial period. The functional sensitivity of 11 selected assays met the clinical requirements (e.g., thyreotroponin (TSH) 0.008 mU/l, troponin T 0.02 microg/l, total prostate-specific antigen (PSA) 0.03 microg/l). The E170 showed no drift during an 8-hour period and no relevant reagent carryover. Accuracy was confirmed by ring trial experiments and method comparisons vs. Elecsys 2010. The reliability and practicability of the system's hardware and software met with, or even exceeded, the evaluator's requirements. Workflow studies showed that E170 can cover the combined workload of various routine analysers in a variety of laboratory environment. Throughput and sample processing time requirements were achieved while personnel 'hands-on-time' could be reduced.

Multicenter analytical performance evaluation of the Elecsys proBNP assay.

The purpose of this multicenter study was to evaluate the technical performance of the automated Elecsys proBNP (brain natriuretic peptide) assay, which is indicated as an aid in the diagnosis of individuals suspected of having congestive heart failure. The Elecsys proBNP assay is an electrochemiluminescent immunoassay employing two polyclonal NT-proBNP-specific antibodies in a sandwich test format. The study was performed on the three Elecsys analyzers (E 1010, E 2010, and E 170) at eight different sites world-wide. Within- and total precision were < or = 3%, with total precision slightly higher on the Elecsys E 170 instrument with multiple modules. Reproducibility among sites and platforms was < 5%. Precision at particularly low NT-proBNP concentrations was assessed down to approximately 25 pg/ml with CVs of 12.6% at 29.2 pg/ml and 9.6% at 38.5 pg/ml for the Elecsys 1010/2010 and E 170, respectively. Linearity was evaluated up to 25,000 pg/ml with a sample-based non-linear response observed with recoveries of < 90% for proBNP concentrations < 10,000 pg/ml. Slopes ranged between 0.92 and 1.02 and intercepts from -5.3 to 10.4 pg/ml (r > or = 0.998) among the three types of analyzers. Slopes were 4.95 and 4.53 in comparison to the Biosite Triage and Shionogi BNP assays. There was no assay interference, and no effect of barrier gels, tube composition, or freeze-thaw. NT-proBNP concentrations in EDTA plasma were up to 10% lower than in serum or heparinized plasma and the analyte was stable at 4 degrees C for up to 72 hours (the maximum time tested). There was no circadian rhythm in normal subjects or congestive heart failure patients and there was no effect of drawing position. In summary, the Elecsys proBNP assay exhibits good technical performance and is suitable for use in routine clinical laboratories to aid in the diagnosis of congestive heart failure.

Multicenter evaluation of analytical performance of the Liaison troponin I assay.

OBJECTIVES: This study evaluated the analytical characteristics of the Liaison immunoassay for cardiac troponin I (cTnI). DESIGN AND METHODS: The protocol consisted of eight sections: evaluation of antibody specificity, linearity, detection limit and imprecision, method comparison, evaluation of endogenous interferents, anticoagulant interference, sample stability, and reference values. RESULTS: The assay equally measured free and complexed cTnI. The minimum detectable cTnI concentration was 0.021 microg/l. The cTnI concentration corresponding to a total CV of 10% was 0.056 microg/l. Linearity of response was demonstrated along the entire dynamic range of the assay. Assay interferences were minimal. cTnI concentrations in serum and heparinized plasma were significantly different. Values in EDTA plasma were on average approximately 5% higher than in matched serum, but this difference was not significant. The 99th percentile cTnI value in healthy subjects was 0.036 microg/l. CONCLUSIONS: Being sensitive, specific, and precise, the Liaison cTnI assay meets current requirements to aid in the diagnosis of myocardial necrosis.

Technical performance and diagnostic utility of the new Elecsys neuron-specific enolase enzyme immunoassay.

This international multicenter study was designed to evaluate the technical performance of the new double-monoclonal, single-step Elecsys neuron-specific enolase (NSE) enzyme immunoassay (EIA) and to assess its utility as a sensitive and specific test for the diagnosis of small-cell lung cancer (SCLC). Intra- and interassay coefficients of variation, determined in five control or serum specimens in six laboratories, ranged from 0.7 to 5.3 (inter-laboratory median: 1.3%) and from 1.3 to 8.5 (inter-laboratory median: 3.4%), respectively. Laboratory-to-laboratory comparability was excellent with respect to recovery and inter-assay coefficients of variation. The test was linear between 0.0 and 320 ng/ml (highest measured concentration). There was a significant correlation between NSE concentrations measured using the Elecsys NSE and the established Cobas Core NSE EIA II in all subjects (n = 723) and in patients with lung cancer (n = 333). However, NSE concentrations were systematically lower (approximately 9%) with the Elecsys NSE than with the comparison test. Based on a specificity of 95% in comparison with the group suffering from benign lung diseases (n = 183), the cut-off value for the discrimination between malignant and benign conditions was set at 21.6 ng/ml. NSE was raised in 73.4% of SCLC patients (n = 188) and was significantly higher (p < 0.01) in extensive (87.8%) as opposed to limited disease (56.7%). NSE was also elevated in 16.0% of the cases with non-small cell lung cancer (NSCLC, n = 374). It is concluded that the Elecsys NSE EIA is a reliable and accurate diagnostic procedure for the measurement of NSE in serum samples. The special merits of this new assay are the wide measuring range (according to manufacturer's declaration up to 370 ng/ml) and a short incubation time of 18 min.