A model for harmonization of routine clinical chemistry results between clinical laboratories.

Clinical chemistry laboratory results from different laboratories often show large between-laboratory variation due to factors such as differences in method principles, method applications, calibration procedures or the application of different instrument factor settings within the same calibration procedure. We have examined the possible use of common calibrators to reduce this variation. Three different calibrators were compared: A, freeze-dried preparations of pooled patients' serum samples, spiked to give three concentration levels; B, freeze-dried preparations of pooled patients' serum samples selected on the basis of elevated enzyme activities at three levels; C, a single calibrator consisting of frozen pooled serum samples. These calibrators were sent to 11 participating laboratories together with 14 fresh patients' serum samples. We report the variation of the results of 21 general clinical chemistry analytes obtained in the patients' serum samples before and after recalculation on the basis of the results of the calibrators. For most analytes the use of a multiple point linear regression calibration function is able to reduce the between-laboratory variation considerably from more than 30% (enzymes) to values well within the bias limits set by European quality specifications, when the necessary conditions are met. These conditions include the commutability of the calibrator(s) with fresh patients' material. For the enzymes, calibrator material originating from selectively pooled patients' samples appeared to be necessary, whereas for the substrates selectively pooled serum calibrators spiked with exogenous supplements may be used. For harmonization to be effective in practice, calibrators need to be stable over time and to carry assigned values set by certified reference laboratories, and the quality performance of participating laboratories should be appropriately monitored.

Evaluation of five different high-density lipoprotein cholesterol assays: the most precise are not the most accurate.

We evaluated the accuracy and performance of four different test kits for the direct determination of high-density lipoprotein (HDL)-cholesterol and compared them with the phosphotungstic acid/MgCl2 assay. All four homogeneous assays were precise (within-run CV of < 2.0% and between-run CV of < 6.4%); both assays based on immuno-inhibition had the lowest CVs (within-run 1.3% and 0.9%; between-run 2.3% and 2.2%). Interference from haemolysis was negligible, but triglyceride concentrations gave a negative interference. The effects of conjugated and unconjugated bilirubin were opposite; conjugated bilirubin showed a negative interference of up to 40%; unconjugated bilirubin interfered positively up to 50%. Using the recently validated indirect phosphotungstic acid/MgCl2 method as a comparison, all four homogeneous assays did not fulfil the National Cholesterol Education Program total error standard, mainly due to the positive biases of 12 to 42%, apparently associated with improper calibrators. Both assays involving immuno-inhibition showed a concentration-dependent bias.

[The magnitude of the 'anion gap']. / De grootte van de 'anion gap'.

The anion gap is used to evaluate disturbances in the acid-base balance, in particular metabolic acidosis. Due to the introduction of new clinical chemical techniques and modern analysers the reference range of the anion gap has changed. Clinical chemical laboratories should establish or verify their own anion gap reference range. Better communication between the laboratory and the clinician with regard to the anion gap is desirable.

Survey of total error of precipitation and homogeneous HDL-cholesterol methods and simultaneous evaluation of lyophilized saccharose-containing candidate reference materials for HDL-cholesterol.

BACKGROUND: Standardization of HDL-cholesterol is needed for risk assessment. We assessed for the first time the accuracy of HDL-cholesterol testing in The Netherlands and evaluated 11 candidate reference materials (CRMs). METHODS: The total error (TE) of HDL-cholesterol measurements was assessed in native human sera by 25 Dutch clinical chemistry laboratories. Concomitantly, the suitability of lyophilized, saccharose-containing CRMs (n = 11) for HDL-cholesterol was evaluated. RESULTS: In the precipitation method group, which included 25 laboratories and four methods, the mean (minimum-maximum) TE was 11.5% (2.7-25.2%), signifying that 18 of 25 laboratories satisfied the TE goal of

Precipitation methods for high-density lipoprotein cholesterol measurement compared, and final evaluation under routine operating conditions of a method with a low sample-to-reagent ratio.

We evaluated six precipitation methods for high-density lipoprotein cholesterol (HDL-chol) determination: the heparin/Mn2+ precipitation reagent method (Hep), two variants of the phosphotungstic acid/Mg2+ method (Tung-L and Tung-B), the dextran sulfate 50,000/Mg2+ method (Dex), the PEG 6000 method (PEG), and the PEG 6000/dextran sulfate 15,000 (PEG/Dex) method. The Tung-B and PEG/Dex precipitation methods have a low sample/precipitation reagent volume ratio (< 0.4). The Tung-B, Dex, PEG, and PEG/Dex methods gave similar values, averaging within 0.1 mmol/L of each other, showing that the precipitation selectivity of these methods is comparable. The precipitation efficiency of Tung-B and Peg/Dex, however, was superior. Ultrafiltration of the supernatants was needed only at triglyceride concentrations > 16.4 mmol/L (undiluted sample) or > 28.0 mmol/L (sample diluted twofold); however, ultrafiltration without dilution was the most accurate method. Results of Tung-B under routine conditions (33 technicians) agreed well with those of the PEG method for 406 normo- and hyperlipidemic plasma samples. By comparison with the HDL-chol method from the Centers for Disease Control and Prevention, the Tung-B method showed a total error of 10.6% which fulfills the criteria of the National Cholesterol Education Program for HDL-chol analysis. In conclusion, with motivated personnel, Tung-B is a reliable, cost-effective method for routine HDL-chol analysis.

"Ocean-to-Ocean Project": a transcontinental external quality assessment trial in clinical chemistry realized throughout Eurasia.

More than 800 diagnostic laboratories situated throughout the Eur-Asian continent--from the Pacific Coast up to the North Sea littoral--were involved in a common survey of External Quality Assessment (EQA). It consisted of the simultaneous measurement of up to 30 analytes of 'general' clinical chemistry using the same batch of control material. The laboratories were associated in four EQA institutions: SKZL (The Netherlands), OQUASTA (Austria), SEKK (Czech Republic) and BKKSystem (Community of Independent States). The results demonstrated the feasibility of such a large-scale survey and provided a realistic idea about the state-of-the-art of laboratory diagnosis in these countries: Besides some local specific problems, such as poor quality of water or the forced use of reagents and calibrators from different sources, there are general problems hindering an efficient process of 'harmonization' in laboratory medicine, namely, the high methodological dispersion especially in the case of enzymes and of some organic analytes. At the same time there is a potential necessity for more concentrated implementation of internal quality assessment into the routine work of laboratories.

Proposed guidelines for the internal quality control of analytical results in the medical laboratory.

The factors involved in analytical quality relate to definition of quality, creation of quality, and control of quality, and errors arise from external and internal sources as well as from permanent and variable factors. Further, the two main types of error are classified as systematic and random errors. Internal quality control (IQC) systems can only operate on the variable factors which are related to batch-to-batch variations (external factors) and to the performance in the laboratory (internal factors). In creating an adequate internal control system, several problems are faced: (i) quality of control materials, (ii) types and frequency of possible errors, (iii) number and types of control materials, (iv) number of replicates of the control, (v) probability of error detection, (vi) probability of false rejection, (vii) consequences of reject signals, (viii) trouble-shooting systems, and (ix) prevention of errors among many other conditions. Gaussian distributions of control results are assumed and the statistical control rules are evaluated in relation to probability of false rejections, Pfr, and probability of error detection, Ped, for the different rules. Combinations of low Pfr and high Ped are obtained by combining results from e.g. four measurements of the same control sample by use of mean and range rules. Further, it is not possible to establish a common control system which can be used for all quantities and analytical procedures; on the contrary, each procedure should have its particular efficient IQC system. These aspects are discussed and a number of guidelines for statistical control rules and problem related internal quality control are presented.

The transferability of a candidate reference method for determination of creatinine in serum.

We developed a candidate reference method for the determination of creatinine in serum. For the acceptance of a reference method it is important that it be rigorously validated against a definitive method and that the method can be transferred from one laboratory to another. This study focussed on the transferability and consisted of two parts: introduction and familiarization with the method in four clinical chemistry laboratories in the Netherlands, followed by independent measurements of Standard Reference Material 909a2 and several commercial quality control materials provided with reference method values according to the protocol of the German Quality Assessment Organisation. The criterion for judging transferability was the mean total error (%) of the five sera used in the accuracy experiment. For creatinine we used a total error of < 2.2%. For Standard Reference Material 909a2 all four laboratories were able to comply with this demand, while only two laboratories met this requirement for the other four sera. The results for the Standard Reference Material 909a2 from the collaborating laboratories demonstrate that this candidate reference method can be successfully transferred without loss of precision and accuracy.

Characterization and classification of external quality assessment schemes (EQA) according to objectives such as evaluation of method and participant bias and standard deviation. External Quality Assessment (EQA) Working Group A on Analytical Goals in Laboratory Medicine.

Within the scope of this paper, the Working Group has attempted to place external quality assessment (EQA) within the whole context of quality management in laboratory medicine. First, the objectives of EQA schemes are defined and current EQA schemes evaluated. In most schemes, the objectives are not defined a priori and do not allow the definition of the origin of unacceptable individual results from participants. There is an ongoing trend for making traditional EQA schemes more interesting for the participants. Analysis of the factors involved in analytical quality allow the definition of the essential analytical tasks of educational EQA schemes. Beside these quality control tasks, educational EQA also includes quality assurance elements. EQA today has not only an important role to play in the assessment of each participant's performance but also in the assessment of the method. Efficiency of the schemes and educational impact can be improved by appropriate scheme designs according to objectives. After this theoretical approach, some practical examples of problem related EQA designs are given.

External quality assessment: currently used criteria for evaluating performance in European countries, and criteria for future harmonization.

A questionnaire was circulated to European countries seeking information on the criteria used for acceptable performance in external quality assessment schemes. Responses were obtained from 21 countries. Fixed limits are used in 13 countries but the basis for these varies widely and includes clinical decision making, biological variation, views of experts, the state-of-the-art, and combinations of approaches. Variable limits based upon statistical analysis of the performance attained are used in 8 countries. The many advantages of harmonization in Europe have prompted the development of criteria based upon within- and between-subject biological variation for use in schemes which circulate single specimen challenges. Currently used criteria, which show much diversity, are compared with these proposals, and the empirical nature of the majority of the former is demonstrated.

Creatinine and surveys: an assessment.

We analysed the results of surveys on creatinine held in The Netherlands during the years 1992, 1993 and 1994. Assay results of 113 samples were reviewed: 88 human sera and 25 samples of animal origin. The results of 5 creatinine assays, 4 based on the Jaffé reaction and 1 enzymatic procedure, are discussed. The enzymatic assay showed by far the best performance, while some of the Jaffé methods differed considerably. All results were evaluated by reference to a HPLC-based selected method for creatinine. Our study shows the need for caution when applying survey performance criteria for creatinine.

Vitamin C and glycohemoglobin.

Three groups of 10 age- and sex-matched nondiabetic volunteers took 0, 750, or 1500 mg of vitamin C each day for 12 weeks. Glycohemoglobin (GHb) was measured by HPLC, electrophoresis, affinity chromatography, and immunoassay at baseline (-4 weeks and -1 day), during supplementation (6 weeks and 12 weeks), and after supplementation ended (6 and 12 weeks). Plasma vitamin C increased twofold during supplementation but, in contrast with the results of Davie et al. (Diabetes 1992; 41:167-73), there were no between-group differences in GHb, glucose, and fructosamine concentrations. Fructosamine may have increased with storage time. The net effects of vitamin C on absolute GHb at 12 weeks vs -1 day (and at 12 weeks vs 12 weeks after) in % GHb amounted to: HPLC -0.035 (-0.050); electrophoresis +0.005 (+0.035); affinity chromatography -0.070 (+0.015); and immunoassay -0.110 (+0.025). We conclude that supplementation of nondiabetics with 750 or 1500 mg of vitamin C daily for 12 weeks does not cause interference in GHb determinations by HPLC, electrophoresis, affinity chromatography, or immunoassay, and does not reduce in vivo Hb glycation.

Testing the accuracy of total cholesterol assays in an external quality-control program. Effect of adding sucrose to lyophilized control sera compared with use of fresh or frozen sera.

We studied the suitability of various types of human serum preparations to test the accuracy of total cholesterol measurements in the External Quality Assessment scheme in The Netherlands, in which approximately 180 laboratories participate. Checked against the certified Abell/Kendall Reference Method, large reagent-dependent negative biases were observed with lyophilized serum that was insufficiently cryoprotected. The biases for the reagents of Du Pont, Roche, and Beckman averaged -16.7%, -9.2%, and -7.6% respectively; the least bias, -0.4%, was obtained with reagent from Boehringer Mannheim. The beneficial effect of cryoprotection with sucrose was demonstrated by the decrease in interreagent variation from 5.4% to 1.9%, the latter value being comparable with the values for fresh and once-frozen pooled serum (1.3% and 1.7%, respectively). We conclude that the detrimental effect of lyophilization on serum matrix can be minimized by suitable cryoprotection with 200 g/L sucrose.

Desirable routine analytical goals for quantities assayed in serum. Discussion paper from the members of the external quality assessment (EQA) Working Group A on analytical goals in laboratory medicine.

The aim of the Working Group was to describe guidelines for deriving desirable analytical goals in laboratory medicine. First, a literature review is given of the different approaches used until now, and some of the most important studies are presented in detail. These approaches are then discussed critically, and the analytical goals proposed by the group are outlined with respect to monitoring and diagnostic testing. The group recommends that, most realistically, analytical quality specifications be biologically based. For diagnostic testing, the aim is achievement of accuracy, allowing the use of common reference intervals when populations are homogeneous for a given quantity. For monitoring (within an individual laboratory and performed with the same instrument), analytical performance should aim at stable operation and low imprecision compared with the within-subject biological variation. Method accuracy is also very important for the comparability of results from different laboratories or instruments.

Between-country comparability of clinical chemistry results: an international quality assessment survey of 17 analytes in six European countries through existing national schemes.

Two lyophilized control sera were distributed through seven national external quality assessment schemes in six European countries--Belgium, Switzerland, France, The Netherlands, Sweden and the United Kingdom--participated in the study. The results for 17 routine analytes were obtained from almost 5000 laboratories for the two sera. The organizers of the schemes were asked to process the results according to a common outlier removal procedure, and submit method-related data if available. The two sera were also distributed through the external/internal scheme of The Netherlands, and the within-laboratory standard deviations calculated in this scheme have been used in a scaling procedure for the external mean values and between-laboratory standard deviations of the participating countries. The results show remarkable agreement in the national mean values for practically all analytes, but considerable differences in the between-laboratory variation. Data from comparable method groups was obtained for 12 analytes from Belgium, France, The Netherlands and the UK. Though revealing some specific differences between methods and countries, the method-related data are generally in agreement with the all-method data. In this study reference method values were only available for cholesterol. The high degree of agreement found suggests, however, that mutual recognition of all-method mean values in national schemes could be acceptable, especially for analytes for which reliable reference methods are not available. The major element of variation is between-laboratory rather than between-country.

[Results of multicenter evaluation of reagents for determination of sodium, potassium and chloride ions using enzyme activation]. / Ergebnisse der multizentrischen Evaluierung von Reagenzien zur Bestimmung von Natrium-, Kalium- und Chloridionen mittels Enzymaktivierung.

We present the results of a multicentre evaluation with Boehringer Mannheim/Hitachi instruments of new "enzymatic" methods for the determination of Na+, K+, and Cl- in serum or plasma. The between-day coefficient of variation was less than 1.4% (Na+), less than 2.6% (K+) and less than 1.7% (Cl-). The linear range of the assays were at least 80 to 200 mmol/l (Na+), 1.5 to 17 mmol/l (K+) and about 30 to at least 200 mmol/l (Cl-). The comparisons with routine flame atomic emission spectrometry and coulometry showed a satisfactory agreement of the test results. The "enzymatic" assays are insensitive to even grossly elevated levels of bilirubin and lipids (sodium, potassium, and chloride assays), NH4+ (potassium assay) and amylase (chloride assay). Interference by various drugs was not detected. Since the new methods can easily be adapted to photometric clinical chemistry instruments, they represent a valuable alternative to the use of ion-selective electrodes, flame atomic emission spectrometry and coulometry.

[Organization of the laboratory and practicability of methods for electrolyte determination]. / Fragen der Organisation des Laboratoriums und der Praktikabilität von Methoden zur Elektrolytbestimmung.

Requirements, possibilities, and pitfalls of electrolyte (sodium, potassium, and chloride) analysis are reviewed within the light of the experiences in the Academic Hospital St. Radboud, Nijmegen, The Netherlands. In view of the ever increasing demands on short turnaround times, attention is paid to problems with specimen delivery, instrumentation and data distribution. The precision levels of available alternatives for electrolyte analysis namely: flame photometry, direct and indirect ion selective electrode methods, dry chemistry, and the newly developed enzymatic approach for sodium and potassium analysis are discussed.