[Introducing the new international vocabulary of metrology]. / Présentation du nouveau Vocabulaire international de métrologie.

The 3rd edition of the "International vocabulary of metrology - Basic and general concepts and associated terms (VIM)" is a fundamental document for the people who are concerned by metrology. This international standard is bilingual (french/english). The VIM contains definitions of metrological concepts, their hierarchy, numerous explicative notes and examples. This approach confirms that the concepts of the VIM are applicable to various disciplines including clinical laboratory sciences.

[Recommendations for metrological control of measuring equipments in the clinical laboratories]. / Recommandations pour la maîtrise métrologique des équipements de mesure au laboratoire d'analyses de biologie médicale (Document D).

The control of the measuring equipment (balances, mass standards, micropipettes, dilutors, volumetric glassware, thermometers...) is essential to obtain reliable analytical results. This control requires knowledge and use of the main standards, instructions for use, metrological verifications and confirmations as well as creation of checking and standardizing certificates ensuring metrological traceability. These items should help to control the quality of the analytical performances (fidelity and trueness in particular).

[Glossary of the acronyms for the institutions concerned by metrology]. / Glossaire des acronymes d'institutions relevant de la métrologie.

Metrology and one of its contributions, metrological traceability represent two fundamental developments for the clinical laboratories. Several international and national institutions take part in these developments. They elaborate recommendations going from concepts to implementation in the clinical laboratories. These activities are helpful for accrediting clinical laboratories.

[Recommendations for expressing uncertainty of measurement of quantitative results in laboratory medicine]. / Recommandations relatives à l'expression de l'incertitude de mesure des résultats quantitatifs en biologie médicale (Document F).

In laboratory medicine, the quantitative results of examinations are interpreted with regard to reference intervals, clinical decision limits or previous results of a patient, from which it is necessary to inform the clinician about the uncertainty of measurement linked to the value of the result. This document explains the problematic of the expression of the uncertainty of measurement. It proposes recommendations concerning a simple way to evaluate uncertainty of measurement using long term internal quality control data and the value of the uncertainty linked to the method calibration. It approaches the determination of analytical goals and the choice of methods and also the comments accompanying the record of results and a help to their interpretation.

[Interference of hemolysis on haptoglobin determined by kinetic immunonephelometry and comparison between phenotypes]. / Interférence de l'hémolyse sur la détermination de l'haptoglobine en immunonéphélémétrie cinétique et comparaison selon les phénotypes.

The aim of the study was to check if a slight and non visible hemolysis to naked eye such as that induced by blood coagulation could interfere in the immunonephelometric measurement of haptoglobin, and if this interference was dependent on the protein phenotype. Results confirmed that blood coagulation induced a non visible hemolysis whose intensity markedly varied from one specimen to another. Under our conditions (kinetic measurement with a Beckman Coulter immunonephelometer), we observed with the sera a negative interference linked to the hemolysis induced by blood coagulation when compared to corresponding plasmas obtained with lithium heparinate (p < 0,005). It was checked also that this anticoagulant did not induce a positive interference. Hemolysis interference was not found phenotype dependent. These results lead us to recommend to perform haptoglobin measurements on heparinised plasmas.

[Influence of pyridoxal phosphate in measuring aminotransferases activities in patients with viral hepatitis]. / Effet du phosphate de pyridoxal dans la mesure des activités aminotransférases chez les patients avec hépatite virale.

Effect of a pyridoxal phosphate (PP) supplementation of reagents used for ALT and AST measurement was studied in serum of 20 patients suffering from viral hepatitis. Measurements of enzyme activities were carried out at 37 degrees C, using an automate (AU 600, Olympus). Significant differences (p < 0.0001) were observed both for ALT and AST, meanwhile they were more marked for ALT than for AST. This difference was associated with a strong interindividual variability regarding PP activation effect on ALT. In conclusion, aminotransferase measurements should be carried out with a reagent supplemented with PP, when the enzyme activity is used to evaluate a cytolysis. The same recommendation applies when ALT results are integrated into various combinations developed for the evaluation of liver status.

A new continuous automated assay for the determination of intestinal lactase.

BACKGROUND: A reliable, sensitive and practicable method for the measurement of intestinal lactase activity is needed. METHOD: The assay is based on the continuous measurement of released glucose by a coupled hexokinase/glucose-6-phosphate dehydrogenase (HK/G6PDH) reagent. RESULTS: The procedure was first optimized for lactase from rat intestinal mucosa. The optimum pH is 6.5 and apparent Km was 17 mmol/l for lactose. The procedure was also adapted on a Cobas Mira automated analyzer; progress curves of the reaction rate can be continuously monitored. The automated assay correlated strongly with the conventional method of Dahlqvist (r(2) = 0.996). The described method has also been applied to human intestinal mucosa biopsies after determination of the catalytic properties of human enzyme (optimum pH 6.0 and apparent Km 34 mmol/l). CONCLUSION: The HK/G6PDH method is reliable, rapid, sensitive and easy to perform both manually as well as in the automated version. It is optimized for human and rat intestinal lactase.

[Enzyme calibrators: principle and practical use]. / Le calibrage en enzymologie clinique: principe, conditions d'application et résultats.

Results of catalytic activities of enzymes are highly dependent on the measurement procedures and on local conditions. Thus, only poorly marked improvement of interlaboratory comparability of results have been observed in clinical enzymology. To solve this problem, SFBC and IFCC have proposed to use "validated enzyme calibrators". Standardised operating procedures adapted to 37 C have been developed by IFCC for the most commonly used enzymes in clinical chemistry, and will be soon published. Reference materials which have been certified with these SOPs can be used as calibrators for a set of measurement methods which exhibit the same analytical specificity. Calibrators must be commutable, a property that must be checked experimentally. It is possible to produce stable and commutable materials for the calibration of a set of methods. Interest of this approach has been demonstrated for several enzymes. Results of two studies presented here show that the comparison of results to the upper limit of reference ranges does not improve the interlaboratory comparability of results in contrast to the calibration of different methods by a common calibrator which allowed to reach an interlaboratory CV close to 4% for ALT and gammaGT.

[Harmonization of practices: application to the measurement of enzymatic activities used in prevention, screening, diagnosis and therapeutic monitoring]. / Harmonisation des pratiques: application à la mesure d'activités enzymatiques utilisées en prévention, dépistage, diagnostic et surveillance thérapeutique.

The large metrological variation (CV, about 25%) observed between laboratories, at the national French level, for the measurement of enzymatic activities results in a loss of efficiency in using laboratory results. Current data show that the standardisation of methods is insufficient to solve this problem and needs to be completed by an harmonisation of the practices including the use of a common reference (calibrator). The present work, carried out by the joint working group between laboratories of the Centres for Periodic Health Examination and the French Society of Clinical Biology (SFBC), deals mainly with the feasibility and evaluation of the improvement of the consistency of the results. Twenty laboratories participated in this study. Five independent surveys were conducted during an height month period. Two enzymes were selected because of their clinical importance and their interest in prevention, screening, diagnosis or epidemiology: ALT (alanine aminotransferase) and GGT (gamma-glutamyltransferase). In each survey three kinds of samples i.e. control sera, candidate calibrators and human serum pools, each of them at two levels of activity (one physiological and the other pathological) were measured in duplicate. The low intra-laboratory imprecision and the high degree of the standardisation of used methods, due to an important effort previously done in this field, permitted to consider a common calibration. The stability and mainly the commutability, i.e. the ability for the candidate calibrator to show a behaviour similar to that of human samples towards the used methods, allowed to reduce the inter-laboratory variation by a half to two third-fold, reaching a coefficient of variation < 5% similar to those observed for cholesterolemia or glycemia. This level of consistency should permit to use common reference limits and common decision limits, after validation of this approach in real practice. The consequences of the harmonisation of practices, extended to the all laboratories, exceed largely the scope of this study. The reduction of the uncertainty and a better approach of the accuracy for the measurement of enzymatic activities should led to a real benefit for the patients in terms of prevention, screening, diagnosis or therapeutic monitoring and consequently for the public health.

Production and certification of an enzyme reference material for adenosine deaminase 1 (BCR 647).

BACKGROUND: We describe the preparation of a lyophilised reference material containing purified human adenosine deaminase 1 and the certification of its catalytic concentration. METHODS: The enzyme was purified from human erythrocytes. RESULTS: The enzyme was >99% pure on polyacrylamide gel electrophoresis. Only trace amounts (<0.4%) of alanine aminotransferase, aspartate aminotransferase and L-lactate dehydrogenase were detected in the purified fraction. The purified adenosine deaminase had a molar mass of 41600 g/mol and an isoelectric pH at 4.7, 4.85 and 5.0. The material was prepared by diluting the purified adenosine deaminase in a matrix containing 50 mmol/l Tris-HCl buffer pH 7.4 and 30 g/l human serum albumin; dispensing in vials and freeze-drying. The batch was homogeneous and the predicted loss of adenosine deaminase activity per year on the basis of accelerated degradation studies was 0.006% at -20 degrees C and 0.04% at 4 degrees C. The certified value for adenosine deaminase catalytic concentration in the reconstituted reference material is (2.55+/-0.09) microkat/l when measured by the method that uses adenosine as substrate and glutamate dehydrogenase as auxiliary enzyme at 37 degrees C. CONCLUSIONS: The material can be used to verify the comparability of results from different laboratories, for intra-laboratory quality control, or for calibration of the adenosine deaminase catalytic concentration measurements.

[C-reactive protein and transthyretin in early diagnosis of infection after open fractures of the lower limbs (a preliminary study)]. / La protéine C-réactive et la transthyrétine dans le diagnostic précoce de l'infection après fracture ouverte des membres inférieurs (étude préliminaire).

PURPOSE OF THE STUDY: The authors investigated the value of C-reactive protein (CRP) and transthyretin (TTR) in the early diagnosis of infection after open fractures of the lower limb in an open, prospective study. MATERIAL AND METHODS: Eighty patients were treated with acute debridement and bone fixation. Follow-up included clinical, radiological, bacteriological and biological assessment: white cell blood count (WBC), erythrocyte sedimentation rate (ESR), CRP and TTR. Diagnosis of infection was based on macroscopic presence of pus. RESULTS: Post-operative reference biological profiles were defined in 74 cases without infection. Reference profiles of WBC and ESR showed unreliable interindividual variations and could not be considered for the diagnosis of infection. Reference profiles of CRP and TTR showed a respective increase (for CRP) and decrease (for TTR) in the early post-operative course, with return to normal values after 12 days. In 6 infected patients, CRP concentrations were suddenly increased and TTR concentrations decreased at the time (3 cases) or even before (3 cases) clinical diagnosis of infection. These variations were mostly simultaneous. No unusual profile was found. The ratio of CRP/TTR concentrations experienced also a sudden increase in infected cases. DISCUSSION: Because of not specifical and unreliable variations in the post-operative outcome of non infected patients, WBC and ESR cannot be considered for the early diagnosis of infection. CRP and TTR concentrations with a respective cut-off value of 100 mg/L and 120 mg/L were found efficient for the early diagnosis of infection, and preceded clinical diagnosis in three of them. A CRP/TTR ratio over 60 p. 100, 8 days or more after initial surgery was found to be very specific (93 p. 100) and sensitive (100 p. 100) for the diagnosis of infection. CONCLUSION: Serial quantifications of CRP and TTR should be performed every four days during the follow-up of open fractures in order to early diagnose a post-operative infection. Comparison of both CRP and TTR could allow a higher accuracy, because of the possible lack of variation of one the two markers.

A continuous automated method for the determination of intestinal disaccharidase activities.

A reliable method for the measurement of various disaccharidase activities such as maltase, isomaltase and sucrase is introduced. It is based on the continuous measurement of liberated glucose by a commercially available glucose dehydrogenase reagent. The procedures were first optimized for enzymes from rat intestinal mucosa. The pH optima were similar (6.3-6.7) for the three enzymes tested, and the apparent Kms were estimated to be 18, 12 and 19 mmol/l for maltase, isomaltase and sucrase, respectively. The procedures were adapted on a Cobas Mira automated analyzer. The assays correlated strongly with the conventional method of Dahlqvist. They were reliable, rapid, easy to perform and validate because the progress curve of each reaction rate can be continuously monitored. The method described has also been applied to human intestinal mucosa (Caco-2 cells).