Development and validation of 14 human serum protein assays on the Roche cobas® c 501

Many laboratories rely on dedicated nephelometers and turbidimeters for the measurement of serum proteins. There are, however, a number of chemistry analyzers that offer open channel configurations for end‐user applications. We developed and validated 14 human serum protein assays (α₁‐antitrypsin, α₂‐macroglobulin, albumin, apolipoproteins AI and B, complement components 3 and 4, haptoglobin, immunoglobulins A, G, and M, orosomucoid, transferrin, and transthyretin) on the Roche cobas^® c 501. We obtained excellent precision at low, normal, and high physiologic concentrations of each protein (within‐run imprecision CVs ≤2.5%, total imprecision CVs ≤3.6%). Linearity for each method was within 5% of the expected value throughout the calibration range, and method comparison studies to commercial assays from Roche or Siemens were in good agreement (r>0.975). We observed no significant interference from bilirubin (up to 414 mg/l), hemoglobin (up to 8.9 g/l), triglyceride (up to 28 g/l), or rheumatoid factor (up to 3,930 IU/ml). Calibration was stable for at least 14 days. The instrument's small reaction cell allowed us to conserve nearly 60% of our specimen and reagent volume compared with our previous system. These newly developed assays provide precise and accurate results with high throughput, but without the associated cost of a dedicated instrument. J. Clin. Lab. Anal. 25:52–60, 2011. © 2011 Wiley‐Liss, Inc.     

Multicenter evaluation of five assays for myoglobin determination

BACKGROUND: Lacking assay standardization, different myoglobin methods may produce results that differ significantly. METHODS: A multicenter study was carried out to compare the analytical performance of five commercially available assays for myoglobin measurement. Linearity, imprecision, interferences, and method comparison were studied according to NCCLS guidelines, whereas reference values were determined following IFCC recommendations. RESULTS: The BNA and Opus showed relatively high imprecision (all but one total CV >7.4%). Other assays showed lower CVs, but they varied among laboratories, particularly at a normal myoglobin concentration (Access, 6.0-11%; Hitachi, 3.8-5.8%; Stratus, 3.4-6.5%). Results were lower in anticoagulated samples on the Access, in heparin and citrate samples on the Stratus, and in citrate samples on the BNA and Opus, and increased in heparin and EDTA samples on the Hitachi. Use of separator gel produced results significantly lower (P <0.001) on the Hitachi and higher (P = 0.016) on the Opus. Bilirubin, turbidity, and hemoglobin had no effect on evaluated methods, but rheumatoid factor affected the Access. In method comparisons, high correlation coefficients (>/=0.98) were obtained. The Stratus gave higher results; however, the Access and BNA gave the lowest. The following upper reference limits (microgram/L) for men and women, respectively, were obtained: Access, 70 and 52; BNA, 51 and 49; Hitachi, 67 and 58; Opus, 80 and 50; and Stratus, 86 and 63. CONCLUSION: The possibility of high imprecision and marked disagreement among commercial myoglobin assays should be carefully considered in clinical practice.     

A multicenter evaluation of the Boehringer Mannheim/Hitachi 717 system

Analytical performance of the Boehringer Mannheim/Hitachi 717 system was evaluated in a multicenter study involving seven different laboratories. Fifty-five methods including end point chemistries, enzymes, ISE, TDM, DAU, and specific protein assays were assessed over a 7 month period. Methods on the analyzer exhibited excellent precision with CVs less than 2% for within run precision, and CVs less than 3% for between day precision for most analytes; linearity, which met or exceeded manufacturer's claims; minimal sample and reagent carryover, and no significant interference from hemolysis; icterus; and lipemia. Recovery of the assigned value for 10 analytes in SRM 909 was acceptable. Comparison of methods with other BM/Hitachi analyzers resulted in slopes close to unity (0.93-1.06); comparison to other clinical chemistry analyzers yielded slopes of 0.88-1.07. Excellent performance and diverse method applications make the BM/Hitachi 717 analyzer a suitable instrument for work station consolidation.     

Precision and comparability of Abuscreen OnLine assays for drugs of abuse screening in urine on Hitachi 917 with other immunochemical tests and with GC/MS

Abuscreen OnLine assays for drugs of abuse screening in urine have recently been developed for use on Hitachi 917 analyzers (Roche Diagnostics GmbH). The assays are based on the kinetic interaction of microparticles as measured by changes in light transmission. Drug in a sample inhibits the formation of particle aggregates and diminishes absorbance change increases. It was the goal of this study to evaluate precision and comparability of the new asssys with CEDIA drugs of abuse tests on Hitachi 917 in different laboratories (three European and three US). The assays were calibrated in the nonlinear mode with four to six standards (semiquantitative application). Initial within-run (21 replicates, four labs) and between-day (10 days, two labs) imprecision studies using Abuscreen OnLine tests and commercial negative (0.5 x cut-off) and positive (1.5 x cut-off) controls revealed the following median CVs [withinrun neg./pos. control/between-day neg./pos. control]: amphetamines 1.9/1.3/3.4/2.4, barbiturates 3.0/1.6/3.9/3.1, benzodiazepines 4.7/1.5/6.3/3.0, cocaine metabolite 1.8/0.9/2.4/1.7, methadone 5.4/1.6/5.5/2.2, opiates 5.5/2.8/5.3/2.7, THC 8.9/4.8/21.8/12.1. CVs < 10% were obtained for the THC test using controls with concentrations closer to the cut-off. An identical set of 170 GC/MS analyzed urine samples was distributed to the six laboratories and measured with Abuscreen OnLine tests on Hitachi 917. The median values for each individual sample were calculated and compared with the results obtained on individual Hitachi 917 analyzers by Passing-Bablok regression analysis. A good agreement between the laboratories was found with less than +/- 11% slope deviation and intercepts below 7% of the cut-off except for benzodiazepines (one slope 17%, one slope--26%) and THC (one slope 34%, one slope--18%). The comparability with CEDIA tests was analyzed by concordance plots using randomized routine samples in three laboratories. The following results were obtained in one of the participating laboratories [cut-off ng/mL] (No. of positive/negative/discrepant samples): amphetamines [500] 2/147/0, barbiturates [200] 1/148/0, benzodiazepines [100] 52/91/7, cocaine metabolite [300] 17/129/3, methadone [300] 113/34/2, opiates [300] 31/114/4, THC [50] 66/81/2. GC/MS was performed for clarification of the discrepant results. In summary, Abuscreen OnLine tests on Hitachi 917 give precise results which compare well when analyzed in different laboratories. They can be rated as convenient and flexible methods for drugs of abuse screening in the routine.     

Clinical vitamin B6 analysis: an interlaboratory comparison of pyridoxal 5'-phosphate measurements in serum

BACKGROUND: Recent investigations into the role vitamin B(6) plays in reducing risk of stroke and cardiovascular disease have heightened interest in vitamin B(6) intake and its relationship to clinical status indicators. Because a true reference method and certified reference materials are lacking, little is known about the relative analytical performance of clinical vitamin B(6) assays. METHODS: Ten laboratories experienced in clinical vitamin B(6) analysis participated in a 3-day analysis of 69 serum and 3 aqueous specimens for pyridoxal 5'-phosphate (PLP). Laboratories used either HPLC-based or enzymatic assays. Results were analyzed for imprecision, recovery, and bias relative to consensus means. RESULTS: Among laboratories, mean within-day CVs (3 specimens x 3 measurements/day) were 0.6%-37% and between-day CVs (20 specimens x 1 measurement/day x 3 days) were 1.4%-26%. Mean recoveries of added PLP were 53%-144%, and mean sample pool mixing recoveries were 75%-119%. Consensus means calculated for 20 serum specimens gave mean relative biases between measurement of -10.0% to 24.3% among participating laboratories over a range of 15.8-319 nmol/L PLP. Measurement imprecision and biases were evaluated against empirically derived performance criteria based on biological variation. Three of 10 laboratories met optimum imprecision requirements and had 90% or more of measurements satisfy optimum criteria for biases among methods. All 10 laboratories met minimum imprecision requirements, but 25%-53% of the results reported by 4 of the 7 suboptimal laboratories failed to satisfy the minimum criteria for bias. CONCLUSION: Agreement among vitamin B(6) methods is good, but large differences in laboratory proficiency exist, pointing to the need for vitamin B(6) reference materials and external quality assurance programs.     

Axon clinical chemistry analyzer evaluated according to ECCLS protocol.

We assessed the analytical performance of the Axon system (Bayer Diagnostici), according to the European Committee for Clinical Laboratory Standards guidelines, for assay of 12 analytes: cholesterol, creatinine, glucose, total protein, urea, uric acid, alkaline phosphatase, alpha-amylase, aspartate aminotransferase, creatine kinase, sodium, and potassium. The field evaluation lasted approximately 5 months and involved the collection of approximately 10,000 data points with the Axon. The following results were obtained: The highest CVs for controls and human sera at different concentration/activity values were 2.2% for within-run imprecision (n = 60; 3 days, pooled estimate) and 3.5% for the between-day imprecision (n = 20 days). Close correlation was found with results for patients' specimens assayed with comparative instruments (Hitachi 717 for substrates and enzymes, Beckman Synchron EL/E4A for electrolytes). No drift was observed during 8 h of operation. The linearity range was broad, sometimes exceeding the manufacturer's claims. No sample-, reagent-, or cuvette-related carryover was found. Measurement of control sera gave results within +/- 5% of the assigned values. We conclude that good reliability and practicability make the Axon system suitable for laboratories with various needs.     

Precision of autoantibody assays in clinical diagnostic laboratories: What is the reality?

BackgroundISO 15189 accreditation remains a challenge for specialized laboratories. In the field of autoimmunity, beside the crucial problem of absence of standardization, laboratories have to manage the analytical performances of the large panel of assays in terms of sensitivity and specificity, but also on their measurement precision for which no reference values are available on biorepositories.MethodsAs an initiative of the French EASI (European Autoimmunity Standardization Initiative) group, French clinical diagnostic laboratories were requested to participate in a survey aiming to analyze the coefficients of variation (CVs) of intra-run and inter-run variability obtained with assays quantifying 14 different autoantibodies. Two performance goals corresponding to the 90th percentile and the 50th percentile (lowest CV values reached by 90% and 50% of laboratories respectively) defined for three levels of concentration were calculated. The impact on the assay performances of the number of measurements, of the nature of the internal quality control (IQC) and the type of immunoassay, was also analyzed.Results414 and 616 values of intra-run and inter-run CVs were collected, respectively. The 50th percentile performance goals were comprised between 1.0% and 8.9% for the intra-run CVs, and between 1.8% and 14.6% for the inter-run CVs. At 90th percentile, the performance goals were comprised between 3.2% and 13.5% for the intra-run CVs, and between 7.3% and 30.8% for the inter-run CVs. CVs calculated from 10 values were similar to those obtained from more values. Higher imprecision was observed when the antibody levels of the IQC was lower than 2 fold the positive threshold. Commercial IQCs gave lower CVs than IQCs derived from patient samples.ConclusionOur results allow proposing some acceptability limits for the precision performances of the autoantibody assays, compatible with the reality of life in diagnostic laboratories and clinical care.     

Evaluation of analytical methods and workflow performance of the Architect ci8200 integrated serum/plasma analyzer system

BACKGROUND: The Architect ci8200 is an integrated serum analyzer for photometric, electrochemical and immunological assays. Several assays of each category and the workflow performance of the system were compared with established laboratory procedures in two laboratories. METHODS: Measurements were compared with the ELECSYS 2010 (Roche Diagnostics) for CEA, PSA, FPSA, AFP, folate, vitamin B12, with the CENTAUR (Bayer) for TSH, T4, FT4, FSH and Estradiol, with the LIAISON (DiaSorin) for TSH, FT4 and FT3, with the Behring Nephelometer BN II (Dade-Behring) for ferritin, and with the INTEGRA 800 (Roche Diagnostics), and the AU640 (Olympus) for clinical chemistry assays. Workflow studies were performed to compare times of analysis required for defined analytical workloads. RESULTS: The coefficients of variation (CVs) for within-run imprecision were between 3% and 6% for CEA, PSA, FPSA, AFP and ferritin, and between 3% and 11% for TSH, FT4, FT3, folate and vitamin B12. The CVs for day-to-day imprecision for immunoassays were between 3% and 10%, except for vitamin B12 (CVs 11-13%) and FT4 (CV 10% -13%). For clinical chemistry tests corresponding CVs for within-run imprecision were < 1%, except for HDL, triglyceride, creatinine, ALT, LD and lipase (CVs<2%) and bicarbonate (CV 3%-6%) and magnesium (CV < 3%). The CVs for day-to-day imprecision for clinical chemistry tests were < 1%, except for sodium, CO(2), magnesium, phosphorus, glucose, uric acid, HDL, triglyceride, ALT, AST CK, lipase with CVs < 6% and for CO(2)<11%. Dilutional linearity testing of seven immunoassays and five clinical chemistry analytes resulted in recovery rates of 90-110%. Correlation studies with 15 immunoassays and 25 clinical chemistry tests showed acceptable agreements with established methods. Work flow analyses demonstrated a net gain in time of analysis up to 109 min depending on the size of the sample batch analyzed with the Architect ci8200 as the main analyzer as compared to the currently installed routine laboratory equipment. Median turn-around times were 7 and 30 min for chemistry assays and immunoassays, respectively, when ordered as STAT analyses, and 18 min when chemistry assays were ordered as routine determinations. CONCLUSIONS: Assays on the Architect ci8200 performed well, fulfilling quality control requirements as defined for instance by German quality control guidelines (RiliBAK). Method comparisons showed acceptable agreements with established assays. Workflow studies using the Architect ci8200 documented shorter times of analyses as compared with the conventionally established laboratory routine demonstrating the potential of integrated chemistry/immunoassay analyzers to provide faster and more efficient performance.     

Automated spectrophotometric analysis of mitochondrial respiratory chain complex enzyme activities in cultured skin fibroblasts

BACKGROUND: Mitochondrial respiratory chain complex (RCC) disorders may occur as commonly as 1 in 8500 individuals. Because of the great variability of phenotypic presentations, measurement of individual RCC enzyme activities is a crucial diagnostic process. Current assay methods are time-consuming and labor-intensive and thus constitute a major impediment to clinical practice. A method with a faster turnaround time would therefore be beneficial. METHOD: We developed an automated spectrophotometric method for measuring the respiratory chain enzyme activities of complex I, complex II + III, and complex IV with the Hitachi 912, an automated spectrophotometer. Mitochondrial citrate synthase was also determined for normalization of the RCC activities. RESULTS: A blinded method comparison with samples from an external testing center yielded a 91% concordance of interpretations. Mean intraassay imprecision (as CV; n = 20) in a single batch analysis of each RCC was 5.9%. Interassay imprecision, evaluated on 2 samples harvested and analyzed 3 times each, gave mean CVs of 10%-18%. CONCLUSIONS: With this automated method, a panel of RCC enzyme activities can be determined in <2 h. In addition, an immunoblot assay using monoclonal antibodies against specific subunits of RCC enzyme complexes can be informative in cases of borderline enzyme activity. Our results suggest that in vitro diagnosis of RCC enzyme deficiencies in skin fibroblasts is an effective alternative to invasive muscle biopsy.     

Performance characteristics of the Architect brain natriuretic peptide (BNP) assay: a two site study

INTRODUCTION: Brain natriuretic peptide (BNP) is produced by the ventricles of the heart and is a biomarker for heart failure. Several commercial assays are now available. We evaluated the performance characteristics of the ARCHITECT BNP assay. METHODS: We evaluated the limit of blank, limit of detection, linearity and imprecision. Method comparison studies were performed with 3 other automated BNP assays including the ADVIA Centaur, AxSYM, and UniCel DxI 800 methods. RESULTS: The mean LOB and LOD of the Architect assay were 3.5 and 5.8 ng/L, respectively. Imprecision studies yielded within run CVs of 1.1 to 5.1% and total CVs of 2.3 to 5.3% using human plasma based multi-constituent controls at concentrations of 92, 500, and 3500 ng/L. The maximum deviation from the target recovery for dilution linearity was 9.6%. Concordance with other BNP assays at a 100 ng/l cutoff was 91 to 98% and kappa statistics were 0.78 to 0.96. The mean difference between the Architect and Advia Centaur methods was positive. For the other methods, the mean difference with the Architect was negative. CONCLUSIONS: The Architect BNP assay shows good performance characteristics with total imprecision < or =5.3%. It agrees well with the Advia Centaur, AxSYM, and UniCel DxI BNP assays.     

The determination of thyroxine and thyroxine uptake with new homogeneous enzyme immunoassays using Boehringer Mannheim/Hitachi analysis systems.

New homogeneous enzyme immunoassays for the determination of thyroxine and thyroxine uptake have been developed. The CEDIA assays are based on the cloned enzyme donor immunoassay technology, which involves fragments of beta-galactosidase prepared by genetic engineering. The assays have been adapted for Boehringer Mannheim/Hitachi analysers. The CEDIA T4/T Uptake assays were evaluated in eleven clinical chemistry laboratories on various Boehringer Mannheim/Hitachi analysis systems, using a 2-point calibration. The analytical range of the T4 test was 10 to 258 nmol/l thyroxine. The T uptake test had a measuring range between 20-50%. Depending on the concentration of the analyte (samples from hypo-, eu- or hyperthyroid patients), mean coefficients of variation ranged from 1.8 to 4.8% within-run and from 4.1 to 6.5% between-run for the T4 assay. Even better coefficients of variation were obtained for the T uptake assay (1.4 to 2.3% within-run, 2.8 to 3.3% between run). The relative inaccuracy of the CEDIA assays with respect to values assigned by other tests was satisfactory in various control sera. The T4 assay was compared with one radioimmunoassay, one enzyme immunoassay and one fluorescence polarisation immunoassay. Slopes ranging from 0.9 to 1.1 and intercepts ranging from -10 to +10 nmol/l thyroxine were obtained with two exceptions. The results of the T uptake test correlated reasonably with those of other thyroxine-binding methods. No interference was observed with icteric and lipaemic sera. Haemoglobin up to 4 g/l had no significant influence. Results of the CEDIA T Uptake test are mainly used for calculation of the free thyroxine index, in which the thyroxine value is corrected for variations of thyroxine-binding protein concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)     

Therapeutic drug monitoring on COBAS INTEGRA 400--evaluation results

The COBAS INTEGRA 400 (Roche Diagnostics GmbH) is a random access analyzer with a consolidated test menue for routine clinical chemistry, specific proteins, drugs of abuse screening and therapeutic drug monitoring (TDM) and different measuring technologies. It was the aim of the present study to evaluate the suitability of this instrument as dedicated analyzer for TDM. Eight assays based on three different technologies were included: Acetaminophen (enzymatic method), Amikacin/ Phenytoin/ Free Phenytoin/ Lidocaine (fluorescence polarization immunoassays; FPIA), Digitoxin/Digoxin (kinetic interaction of microparticles in solution; KIMS). The study comprised the determination of imprecision according to NCCLS EP-T protocol, method comparison and linearity studies. The assays were compared with the corresponding methods on AxSYM or TDx analyzers (Abbott Laboratories). For Acetaminophen and Amikacin COBAS INTEGRA 700 was used as additional comparison instrument. The results are summarized in a table (table 6). Precision results are well acceptable with within-run CVs < 5% and total CVs < 6% except for Digitoxin and Digoxin which show a somewhat higher imprecision at low concentrations. Results obtained for Acetaminophen and Amikacin on COBAS INTEGRA 400 and 700 show excellent agreement. A good comparability is also found between COBAS INTEGRA 400 and AxSYM or TDx methods with slight systematic deviations for Acetaminophen, Amikacin and Free Phenytoin. The lower correlation coefficient for the digoxin method comparison can be attributed to two discrepant samples. Linearity throughout the range studied which covered > 80% of the measuring range was confirmed for the five assays tested (Digitoxin, Digoxin, Lidocaine, Free Phenytoin, Phenytoin) based on the acceptance criteria of +/- 10% deviation of the measured values from the theoretical values. Based on the analytical performance of the TDM tests studied it can be concluded that the COBAS INTEGRA 400 is very well suited for routine TDM analysis.     

Standardization of C-peptide measurements

BACKGROUND: C-peptide is a marker of insulin secretion in diabetic patients. We assessed within- and between-laboratory imprecision of C-peptide assays and determined whether serum calibrators with values assigned by mass spectrometry could be used to harmonize C-peptide results. METHODS: We sent 40 different serum samples to 15 laboratories, which used 9 different routine C-peptide assay methods. We also sent matched plasma samples to another laboratory for C-peptide analysis with a reference mass spectrometry method. Each laboratory analyzed 8 of these samples in duplicate on each of 4 days to evaluate within- and between-day imprecision. The same 8 samples were also used to normalize the results for the remaining samples to the mass spectrometry reference method. RESULTS: Within- and between-run CVs ranged from <2% to >10% and from <2% to >18%, respectively. Normalizing the results with serum samples significantly improved the comparability among laboratories and methods. After normalization, the differences among laboratories in mean response were no longer statistically significant (P = 0.24), with least-squares means of 0.93-1.02. CONCLUSIONS: C-peptide results generated by different methods and laboratories do not always agree, especially at higher C-peptide concentrations. Within-laboratory imprecision also varied, with some methods giving much more consistent results than others. These data show that calibrating C-peptide measurement to a reference method can increase comparability between laboratories.     

Evaluation of an assay for serum 1,5-anhydroglucitol (GlycoMark) and determination of reference intervals on the Hitachi 917 analyzer

BACKGROUND: 1,5-Anhydroglucitol (1,5-AG) is a glucose analogue, which is decreased in hyperglycemic individuals. We report the technical performance of an assay (GlycoMark) on a chemistry analyzer, evaluation of analyte stability and determination of reference intervals for 1,5-AG in a non-diabetic US population. METHODS: NCCLS protocols were followed to evaluate the reagent on a Hitachi 917 chemistry analyzer. RESULTS: Intra- and interassay imprecision ranged from 1.3% to 3.8% and 0.79% to 3.7%, respectively. The assay was linear to 110 microg/ml. Interference from triglyceride, hemoglobin and bilirubin was <10% to concentrations of 12.6 mmol/l, 12.1 and 911.4 micromol/l, respectively. Correlation coefficients between lot numbers on the Hitachi 917 and between analyses on the Hitachi 917 and the Hitachi 7170 analyzers were >0.99. The lowest limit of detection was 0.49 microg/ml (mean+/-2 S.D.). 1,5-AG was stable at 4 degrees C for 7 days, at 22 degrees C for 5 days, at -80 degrees C for 14 days and for three freeze-thaw cycles at -80 degrees C. The US reference intervals (nonparametric 2.5th-97.5th percentiles) were 10.2-33.8 microg/ml (males) and 5.9-31.8 microg/ml (females). CONCLUSIONS: The performance of the GlycoMark assay for the measurement of 1,5-AG was acceptable on the Hitachi 917 analyzer.     

Inter-laboratory evaluation of the COBAS INTEGRA 400 analytical system.

COBAS INTEGRA 400 is a random-access analytical system consolidating assays for clinical chemistry analytes, electrolytes, serum proteins, drugs of abuse and therapeutic drugs. Analytical performance and practicability of the instrument were evaluated in seven laboratories over a 2-year period in parallel with system development. Good within-run and total imprecision for all assays was observed with a few exceptions for specimen pools with low concentration or activity. The coefficients of variation for total imprecision were well below 3.0% for clinical chemistry analytes and electrolytes, and below 5.0% for serum proteins and therapeutic drugs. Method comparisons demonstrated a good agreement with the various systems used for comparison, with slopes varying typically from 0.94 to 1.05, and Spearman correlation coefficient generally > 0.975. Accuracy was verified by recovery of controls and certified reference materials within 90 to 110% of target values. Assay ranges were linear within +/- 5%. No carry-over on reagent or sample pipetting systems was observed. Manufacturer-specified interference limits and onboard stabilities of reagents were confirmed. A time study for calculating direct personnel times and total processing time was carried out in three laboratories under different conditions including consolidated, STAT and dedicated use. On a scenario-independent basis, the total working time was shorter on the COBAS INTEGRA 400 than on routine systems in all three laboratories. Personnel time, in particular, was significantly reduced when compared to routine instruments. In general, system practicability was judged very positively in all laboratories. Owing to its versatility, the instrument is best placed as a consolidated workstation in small- to medium-sized laboratories or as an instrument for special determinations such as serum proteins, drugs, urinalysis or emergency analyses in large laboratories.     

Determination of total and free plasma carnitine concentrations on the Dade Behring Dimension RxL: integrated chemistry system

BACKGROUND: L-carnitine is a naturally occurring quaternary ammonium compound present in all mammalian species. Its major function is to facilitate the passage of long-chain fatty acids through the mitochondrial membrane for subsequent beta-oxidation and ketone synthesis. Clinical interest in carnitine disorders relates particularly to possible deficiency states that may result in a phenotypic spectrum that includes cardiomyopathy, skeletal myopathy, hypoglycemia and hyperammonemia. The objective of this study was to develop a method on the Dade Behring Dimension RxL analyzer for measuring free and total carnitine levels in plasma. METHODS: Plasma samples were deproteinized by ultrafiltration to remove interference by endogenous thiols. Filtrates were measured directly on the RxL for free carnitine or after alkaline hydrolysis for total carnitine by an endpoint enzymatic assay that uses carnitine acetyltransferase. RESULTS: Within-run imprecision was <5% at high and low levels for both free and total carnitine while between-day imprecision was <15%. Recovery of free carnitine from spiked plasma >90%. The method was linear between 5.0 and 150.0 micromol/l and the limit of quantification was 5.0 micromol/l. Comparison of our method with another automated procedure developed on the Hitachi 917 system using Deming regression analysis resulted in the following equations: Dimension=1.034(Hitachi)-7.44 for total carnitine (r=0.955) and Dimension=0.805(Hitachi)+1.96 for free carnitine (r=0.951), respectively. CONCLUSIONS: Our method is suitable for analyzer platforms where the level of imprecision is lower and the throughput is higher than manual methods. It also avoids the use of radioisotopes and is appropriate in labs where access to reference methods such as tandem mass spectrometry and HPLC is limited or unavailable.     

A new liquid homogeneous assay for HDL cholesterol determination evaluated in seven laboratories in Europe and the United States.

We evaluated a new liquid homogeneous assay for the direct measurement of high density lipoprotein cholesterol (HDL-C Plus) in seven laboratories. The assay includes two reagents which can be readily used in most available clinical chemistry analyzers. The total CVs of the new method were below 4.6% and the bias in relation to the designated comparison method was below 3.9%. The total error ranged between 4 to 7%. HDL-C values determined by this method were in good agreement with those obtained by the old homogeneous assay using lyophilized reagents, and other homogeneous and precipitation assays (0.944 < r < 0.996). The assay was linear up to at least 3.89 mmol/l HDL-C. Hemoglobin did not interfere, whereas in icteric samples slight deviations were observed. Lipemia up to 11.3 to 22.6 mmol/l triglycerides did not interfere with this homogeneous HDL-C assay. In samples of patients with paraproteinemia, discrepant results were seen. This liquid homogeneous HDL-C assay was easy to handle and produced similar results in all laboratories participating in this study. This method will enable clinical laboratories to reliably measure HDL-C for risk assessment of coronary heart disease.     

Evaluation of an enzymatic homocysteine assay for the Hitachi series chemistry analyzer

BACKGROUND: Homocysteine is an essential amino acid, whose clinical utilization includes detection of homocystinuria, vitamin B12 and folate deficiencies and cardiovascular disease management. We report assay performance of the Catch reagents on the Hitachi 917 chemistry analyzer. METHODS: Linearity, recovery, imprecision and interference from lipids, bilirubin, hemoglobin and ascorbic acid were determined on a Hitachi 917 chemistry analyzer. Split sample aliquots were assayed using the described method, by HPLC and fluorescence polarization immunoassay for method comparisons. RESULTS: The assay was linear to at least 45 micromol/l. Intra- and interassay variation ranged from 1.7% to 3.4% and 3.3% to 6.7%, respectively. Interference was observed when hemoglobin concentrations was >7 g/l and intralipid >5 g/l. No interference was observed from bilirubin or ascorbic acid. The assay was in good agreement with values obtained by HPLC and fluorescence polarization immunoassay. CONCLUSIONS: The performance of the Catch homocysteine assay was acceptable on the Hitachi 917 chemistry analyzer.     

不同仪器检测肌酐的线性调查

目的 调查不同仪器检测肌酐的线性,旨在评价临床实验室检测肌酐的能力.方法 利用美国临床和实验室标准协会(Clinical and Laboratory Standards Institute,CLSI)推荐的EP6-A指南,即多项式回归分析方法判断统计学标准的线性和非线性,利用Kroll医生对EP6-A补充的方法计算数据的不精密度和最优拟合曲线与直线的平均偏离值.按照分析仪器的不同将检测数据分为4组,即Beckman LX组(28家实验室)、Beckman CX组(14家实验室)、Hitachi组(62家实验室)和Olympus组(72家实验室).结果 在Beckman LX组、Beckman CX组、Hitachi组和Olympus组,肌酐检测数据的不精密度范围分别是0.30%～3.01%、0.09%～3.46%、0.14%～4.91%和0.17%～16.44%,在Olympus组,有1家实验室检测数据的不精密度(16.44%)超过了设定值,其他实验室的检测数据具有作线性调查的精密度.最优拟合曲线与直线的平均偏离值的范围分别是0%～2.38%、0%～2.51%、0%～5.46%和0%～4.66%,统计学标准的线性分别占21.4%(6/28)、35.7%(5/14)、11.3%(7/62)和18.1%(13/72),临床标准的线性分别占78.6%(22/28)、64.3%(9/14)、88.7%(55/62)和80.6%(58/72).通过肌酐线性调查的实验室分别占100%(28/28)、100%(14/14)、100%(62/62)和98.6%(71/72).结论 参加线性调查的实验室大部分具有检测肌酐的能力.     

A multicentre evaluation of the Boehringer Mannheim/Hitachi 704 analysis system

The selective multitest Boehringer Mannheim/Hitachi 704 analysis system was examined according to the ECCLS guidelines in a multicentre evaluation involving four laboratories. Ten routine parameters, covering most of the application settings of the instrument, were measured in the respective laboratory at temperatures 25, 30 or 37 degrees C. The trial lasted four months and gave more than 40,000 data. It yielded the following results: 1. Within the four laboratories the mean coefficients of variation for three control sera at different concentrations were found to be equal to or better than 1.6% for the within-run imprecision and 2.8% or better for the between-day imprecision. 2. No drift was observed during eight hours. 3. Because of the high linear measuring range a re-run analysis was seldom necessary. 4. Sample-related carry-over was not seen. Reagent-dependent carry-over was measured from cholesterol to uric acid and from triacylglycerols to lipase. Through modification of the cholesterol and triacylglycerol reagents, the carry-over effect was practically eliminated. 5. The recovery of the assigned values of control sera showed average values between 99 and 104%. For bilirubin, creatinine, creatine kinase and alanine aminotransferase some control sera showed deviations greater than 10%. 6. In all cases, regression analysis of the results obtained in comparisons of the present instrument with the Hitachi 705 or 737 yielded slopes close to unity with extreme values of 0.95 and 1.06. 7. During the entire evaluation period there was no malfunction or breakdown of the instruments. The evaluators came to the conclusion that the analytical performance as well as the reliability and practicability of the Hitachi 704 can be rated as excellent.