Critical comments to a recent EFLM recommendation for the review of reference intervals.

In a recent EFLM recommendation on reference intervals by Henny et al., the direct approach for determining reference intervals was proposed as the only presently accepted "gold" standard. Some essential drawbacks of the direct approach were not sufficiently emphasized, such as unacceptably wide confidence limits due to the limited number of observations claimed and the practical usability for only a limited age range. Indirect procedures avoid these disadvantages of the direct approach. Furthermore, indirect approaches are well suited for reference limits with large variations during lifetime and for common reference limits.

Multicentre evaluation of a second generation point-of-care assay with an extended range for the determination of N-terminal pro-brain natriuretic peptide.

BACKGROUND: In the second generation of the point-of-care (POC) assay Roche CARDIAC proBNP, the upper limit of the measuring range was extended from 3000 to 9000 ng/L. METHODS: A thirteen-site multicentre evaluation was carried out to assess the analytical performance of the POC NT-proBNP assay and to compare it with a laboratory N-terminal pro-brain natriuretic peptide (NT-proBNP) assay. RESULTS: In method comparisons of six lots of POC NT-proBNP with the lab reference method (Elecsys proBNP) mean bias ranged from -10 to +17%. In lot-to-lot comparisons all six investigated lots of POC NT-proBNP showed excellent agreement, with mean bias between -7% and +2%. The majority of all coefficients of variation obtained from ten-fold measurements using 56 native blood samples were below 8%. No interference was observed with hemolytic blood (hemoglobin concentrations up to 0.12 mmol/L), lipemic blood (triglyceride concentrations up to 14.0 mmol/L) nor icteric blood (bilirubin concentrations up to 63 micromol/L). Hematocrit values between 24% and 51% had no influence on the assay result. High NT-proBNP concentrations above the measuring range of POC NT-proBNP did not lead to false low results due to potential high-dose hook effect. Results with POC NT-proBNP were not influenced by different ambient temperatures (18 degrees C to 32 degrees C), the sample material used, nor by over- or underdosing by 15 microL compared to the regular sample volume of 150 microL. CONCLUSIONS: The POC NT-proBNP assay showed an excellent analytical performance including a good agreement with the laboratory method. The assay is therefore suitable for its intended use in point-of-care settings.

Development and calibration of a new point-of-care test for the determination of NT-proBNP in whole blood.

A new point-of-care test for the determination of NT-proBNP in whole blood was developed based on the existing gold-label rapid immunoassay technology of the Roche Cardiac reader system. The novel gold-labelled monoclonal antibody recognizes NT-proBNP at amino acid sequence 27 to 31, the biotinylated polyclonal antibody recognizes sequence 39 to 50. In a model assay based upon the reference method Elecsys proBNP and with an R & D lot of the point-of-care test, this newly selected and developed combination of antibodies showed a very good correlation with the standard Elecsys proBNP assay with correlations of 0.96 or 0.94, respectively. The test was calibrated according to the existing masterlot concept of the Roche CARDIAC tests with Elecsys proBNP as a reference. In a preliminary method comparison with Elecsys proBNP the accuracy of the calibration was confirmed; the bias was between 1 and 6%. Possible reasons of approximately 1% outliers (> +/- 100%) were discussed.

A plea for intra-laboratory reference limits. Part 2. A bimodal retrospective concept for determining reference limits from intra-laboratory databases demonstrated by catalytic activity concentrations of enzymes.

BACKGROUND: The current recommendations for establishing intra-laboratory reference limits (RLs) cannot be fulfilled by most laboratories because of the expense involved. In the current study, a bimodal method was developed to derive RLs from data stored in a laboratory information system without any assumption concerning the distribution of the diseased subgroup. METHODS: A smoothed kernel density function (D(mix)) was estimated for the distribution of combined data for non-diseased and diseased adult subjects. It was assumed that the "central" part of the distribution represents the non-diseased population, which was defined and used to estimate a Gaussian distribution of either the original values or Box-Cox transformed data. This normal distribution was now considered the distribution of the non-diseased subgroup (D(nd)). Percentiles were calculated to obtain retrospective RLs. The density function of the diseased subgroup (D(d)) was calculated by subtracting the non-diseased density function from D(mix) (D(d)=D(mix)-D(nd)). The intersection point of the D(nd) and D(d) curves identified the RL with the highest diagnostic efficiency. RESULTS: The model was applied to catalytic activity concentrations of several enzymes with data from different laboratories. The RLs obtained were similar to recently published consensus values. Differences between laboratories were small but significant. Gender stratification was necessary for alanine aminotransferase (ALT), aspartate aminotransferase (AST), and gamma-glutymaltransferse (gamma-GT), not significant for lipase and amylase and inconsistent among the laboratories for alkaline phosphatase (AP) and lactate dehydrogenase (LDH). Age stratification was only tested for two groups (18-49 and >or=50 years) and was significant for AST (females only), gamma-GT and lipase, not significant for amylase and inconsistent for AP, LDH and ALT. For gamma-GT, further stratification for age in decades was necessary for males. Creatine kinase MB (CK-MB) values were not stratified owing to the low number of data available. CONCLUSIONS: Retrospective RLs derived from intra-laboratory data pools for the catalytic activity concentration of enzymes using a modified procedure plausibly agreed with published consensus values. However, most RLs varied significantly among laboratories, thus supporting the "old" plea for intra-laboratory RLs.

Multicentre evaluation of a new point-of-care test for the determination of NT-proBNP in whole blood.

BACKGROUND: The Roche CARDIAC proBNP point-of-care (POC) test is the first test intended for the quantitative determination of N-terminal pro-brain natriuretic peptide (NT-proBNP) in whole blood as an aid in the diagnosis of suspected congestive heart failure, in the monitoring of patients with compensated left-ventricular dysfunction and in the risk stratification of patients with acute coronary syndromes. METHODS: A multicentre evaluation was carried out to assess the analytical performance of the POC NT-proBNP test at seven different sites. RESULTS: The majority of all coefficients of variation (CVs) obtained for within-series imprecision using native blood samples was below 10% for both 52 samples measured ten times and for 674 samples measured in duplicate. Using quality control material, the majority of CV values for day-to-day imprecision were below 14% for the low control level and below 13% for the high control level. In method comparisons for four lots of the POC NT-proBNP test with the laboratory reference method (Elecsys proBNP), the slope ranged from 0.93 to 1.10 and the intercept ranged from 1.8 to 6.9. The bias found between venous and arterial blood with the POC NT-proBNP method was < or =5%. All four lots of the POC NT-proBNP test investigated showed excellent agreement, with mean differences of between -5% and +4%. No significant interference was observed with lipaemic blood (triglyceride concentrations up to 6.3 mmol/L), icteric blood (bilirubin concentrations up to 582 micromol/L), haemolytic blood (haemoglobin concentrations up to 62 mg/L), biotin (up to 10 mg/L), rheumatoid factor (up to 42 IU/mL), or with 50 out of 52 standard or cardiological drugs in therapeutic concentrations. With bisoprolol and BNP, somewhat higher bias in the low NT-proBNP concentration range (<175 ng/L) was found. Haematocrit values between 28% and 58% had no influence on the test result. Interference may be caused by human anti-mouse antibodies (HAMA) types 1 and 2. No significant influence on the results with POC NT-proBNP was found using volumes of 140-165 muL. High NT-proBNP concentrations above the measuring range of the POC NT-proBNP test did not lead to false low results due to a potential high-dose hook effect. CONCLUSIONS: The POC NT-proBNP test showed good analytical performance and excellent agreement with the laboratory method. The POC NT-proBNP assay is therefore suitable in the POC setting.

Comparison of cardiac troponin T and I in healthy men and in aortic valve replacement.

Troponins are of outstanding importance for the diagnosis of myocardial infarction. Cardiac troponin T (cTnT) and the various cardiac troponin I (cTnI) assays differ with respect to method comparison, diagnostic sensitivity and diagnostic specificity. To understand the differences in the diagnostic behavior of troponin assays, AccuTnI and Elecsys Troponin STAT were used in a group of healthy men and in the follow-up of patients with aortic valve replacement (AVR). Within the healthy subjects AccuTnI was able to differentiate two subgroups from each other, whereas the cTnT concentrations of all subjects were below the detection limit. In AVR patients, cTnT and cTnI correlated sufficiently, if the postoperative periods were taken into consideration. There was a rapid increase in cTnI within 24 h. In contrast, a broad peak was evident for cTnT between 48 and 120 h. The results emphasize more the differences in the release of cTnI and cTnT from the cytoplasm and the thin filaments of the cardiomyocytes than the modifications of the troponins circulating in the blood.

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.

The European Register for Specialists in Clinical Chemistry and Laboratory Medicine: guide to the Register Version 2-2003 and procedure for re-registration.

The European Communities Confederation of Clinical Chemistry and Laboratory Medicine (EC4) opened a Register for European Chemists in 1997. The operation of the Register is undertaken by a Register Committee (EC4RC). During the last 5 years more than 1,400 clinical chemists entered the register. In this article an update of the first Guide to the Register is given, based on the experience of 5 years of operation and the development of the discipline. The registration is valid for 5 years. In a second part the procedure and the conditions for re-registration are presented.

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.

The practice of clinical chemistry in the European Union.

The European Communities Confederation of Clinical Chemistry has been actively engaged in raising the level of clinical chemistry in the European Union. Closer contacts between the national societies for clinical chemistry have resulted in more comparable programs for postgraduate training of clinical chemists, closer similarity of contents and practice of the profession in the different countries, and the official registration of professionals. This article reviews some of the characteristics of professional organisation, practice, and regulation in the fifteen European Union countries. Many similarities appear. In half of the countries microbiology, blood-banking and transfusion medicine fall within the domain of clinical chemistry. The minimum number of years for training (university and postgraduate) is eight, but in practice this will extend to 10 or more years. Official regulation of the profession by law exists in a minority of countries. Continuing education and re-registration have not been officially instituted yet in any country, but these issues will be the next steps forward. In those countries that prepare themselves for entering the European Union, training and practice of clinical chemistry are moving towards the common standards of the European Communities Confederation of Clinical Chemistry.