Laboratoriumgeneeskunde: van evolutie naar revolutie

Oratie uitgesproken door Prof.dr. C.M. Cobbaert bij de aanvaarding van het ambt van hoogleraar in de Klinische Chemie en Laboratoriumgeneeskunde aan de Universiteit Leiden op vrijdag 19 februari 2016Oratie uitgesproken door Prof.dr. C.M. Cobbaert bij de aanvaarding van het ambt van hoogleraar in de Klinische Chemie en Laboratoriumgeneeskunde aan de Universiteit Leiden op vrijdag 19 februari 2016

Regional serum cholesterol differences in Belgium: do genetically determined cardiovascular risk factors contribute?

BACKGROUND: Differences in serum lipid distribution and mortality from ischaemic heart disease have repeatedly been reported between Belgian northerners and southerners. We investigated whether serum lipoprotein(a) (Lp(a)) and apolipoprotein (apo) E polymorphism were involved. METHODS: Fasting serum lipids, apo A-I and B, and Lp(a) levels were examined in randomly selected, 20-39 year old Belgian males and females from the north (Flanders) and the south (Wallonia) of Belgium (N = 900). Apo E phenotype distribution was investigated in random subsamples from either region (N = 249). RESULTS: Mean serum cholesterol, low density lipoprotein cholesterol (LDL-c), apo B and triglyceride levels were higher in Walloons compared to Flemings within each gender, the difference being significant in 30-39 year old males. Average high density lipoprotein cholesterol and apo A-I levels were significantly lower in 30-39 year old male southerners, compared to their northern counterparts. Median Lp(a) was 67 mg/l in northerners and 75 mg/l in southerners (NS). The apo E phenotype distribution was similar in both regions (chi2 = 7.213; d.f. = 5; P = 0.2053), whereas the average effects of the apo E alleles differed between the regions. In southerners the epsilon4 effect upon adjusted apo B and LDL-c levels was approximately+12% and the epsilon2 effect was approximately-15%; in northerners the epsilon4 and epsilon2 effects were approximately+5% and approximately-25%, respectively. The apo E polymorphism did not affect serum Lp(a) levels. CONCLUSIONS: Regional cholesterol differences between Flemings and Walloons cannot be explained by differences in serum Lp(a) or apo E phenotype distribution. The less favourable epsilon2 and epsilon4 effects in southerners compared to northerners reflect modulation of the apo E gene by particular environments

Quantifying apolipoprotein(a) in the era of proteoforms and precision medicine

Lipoprotein(a) (Lp(a)) is an independent risk factor in the development of atherosclerotic cardiovascular diseases (ASCVD) and calcific aortic valve disease (CAVD). Lp(a) is an LDL-like particle to which apolipoprotein (a) (apo(a)) is covalently bound. Apo(a) contains a variable number of kringle IV repeats, a kringle V and a protease domain. Serum/plasma Lp(a) concentrations are traditionally expressed as total particle mass in mg/L. Concern has arisen lately as flawed Lp(a) mass tests have masked its clinical utility.The determinants of variability in Lp(a) composition were investigated, including the apo(a) size polymorphism, post-translational modifications -N- and O-glycosylation- and the lipid:protein ratio. Depending on the number of kringle IV-2 repeats, the theoretical protein content of the Lp(a) particle varies between 30 and 46 (w/w) %, which inescapably confounds Lp(a) mass measurements.The authors advocate that reporting of Lp(a) particle concentrations in mass units is metrologically inappropriate and should be abandoned, as it results in systematically biased Lp(a) results. Enabling technology, such as mass spectrometry, allows unequivocal molecular characterization of the apo(a) measurand(s) and accurate quantitation of apo(a) in molar units, unaffected by apo(a) size polymorphism. To guarantee that Lp(a)/apo(a) tests are fit-for-clinical-purpose, basic metrology principles should be implemented upfront during test development.Afdeling Klinische Chemie en Laboratoriumgeneeskunde (AKCL

Effect of the reaction temperature on the prothrombin time and the apparent International Normalized Ratio determined with International Standards for thromboplastins

Introduction The definition of the International Normalized Ratio (INR) depends on a reference measurement procedure for the prothrombin time (PT) determined with international standards for thromboplastins. The agreed water bath temperature for PT determination in the reference measurement procedure is 37 degrees C. The aim of the study was to assess the influence of small deviations of the agreed reaction temperature on PT and INR determined with World Health Organization international standards for thromboplastins rTF/16 (recombinant human) and RBT/16 (rabbit brain). Methods Prothrombin time was determined, with a manual hook technique, in glass test tubes in a water bath at a controlled temperature. The PT reaction temperatures were varied between 28 and 40 degrees C. Pooled normal plasma and pooled coumarin plasma (INR approximate to 2.8) were used as test plasmas. The data were fitted to a quadratic relationship between PT and temperature. Results Prothrombin times with rTF/16 were shortened by increasing the reaction temperature up to approximately 39-40 degrees C. PTs with RBT/16 were shortened by increasing the reaction temperature up to approximately 34-37 degrees C, but were prolonged at higher temperatures. The apparent INR change of the coumarin plasma at 37.0 degrees C was 0.06/degrees C and 0.11/degrees C for rTF/16 and RBT/16, respectively. Conclusions Reaction temperature had a significant effect on PT and the apparent INR with the International Standards. At 37.0 degrees C, the apparent INR of coumarin plasma determined with RBT/16 was more responsive to temperature change than the apparent INR determined with rTF/16. The required accuracy of the water bath temperature should be 37.0 +/- 0.1 degrees C.Afdeling Klinische Chemie en Laboratoriumgeneeskunde (AKCL

Significance of various parameters derived from biological variability of lipoprotein(a), homocysteine, cysteine, and total antioxidant status

Analytical and biological components of variability and various derived indices have been determined for lipoprotein(a) [Lp(a)], homocysteine (Hcy), cysteine (Cys), and total antioxidant status (TAOS) in ostensibly healthy adult Caucasians and in stable outpatients with an increased serum Lp(a). In healthy Caucasians, average intraindividual biological CVs (CVb) were 20.0% for Lp(a), 9.4% for Hcy, 5.9% for Cys, and 2.8% for TAOS, CVbs being similar in men and women. In the outpatient group, CVbs were comparable for Hcy, Cys, and TAOS, but significantly lower for Lp(a) (7.5% vs 20.0%; P <0.0001). Moreover, a significant inverse relation between both biological and analytical CVs (CVa) and serum Lp(a) concentrations was demonstrated. We conclude that average CVa and CVb values, and hence average derived indices, are adequate for Hcy, Cys, and TAOS, whereas individual values should be used for Lp(a)

The quest for equivalence of test results: the pilgrimage of the Dutch Calibration 2.000 program for metrological traceability

Afdeling Klinische Chemie en Laboratoriumgeneeskunde (AKCL

Should LC-MS/MS be the reference measurement procedure to determine protein concentrations in human samples?

Afdeling Klinische Chemie en Laboratoriumgeneeskunde (AKCL

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 </=13% issued by the National Cholesterol Education Program (NCEP). In the homogeneous HDL-cholesterol method group, which included five laboratories, each performing two different methods, the mean (minimum-maximum) TE was 9.5% (6.0-17.3%) for the Boehringer assay and 15.7% (3.3-30.7%) for the Genzyme assay. For the Boehringer homogeneous assay, one of five laboratories did not meet the TE criterion, whereas for the Genzyme homogeneous assay, three of five laboratories exceeded the 13% criterion. The biases on the HDL-cholesterol values found by various precipitation methods were highly variable in all CRMs, irrespective of the quality, whereas the biases found by the homogeneous method from Boehringer were far less than +/-5% for the highest-quality CRMs (CRMs 4-6). CONCLUSIONS: The NCEP goal was met by 24 of 35 laboratories assessed by use of native human sera. Selectively pooled, lyophilized CRMs that are cryoprotected with 200 g/L saccharose have ample potential for use in the standardization of homogeneous HDL-cholesterol methods

Noninvasive assessment of reperfusion and reocclusion after thrombolysis in acute myocardial infarction.

The clinical significance of ST-segment changes and of the time course of appearance in serum of different cardiac proteins has been reviewed for the diagnosis of coronary reperfusion and reocclusion after thrombolysis. In particular, the value of serial 12-lead electrocardiographic (ECG) studies, of Holter monitoring, and of continuous multilead computer-assisted ECG monitoring is compared. Regarding the serum proteins, the clinical significance of reperfusion indices described so far for serum creatine kinase (CK), its isoenzyme serum creatinine kinase MB, the CK isoforms, and myoglobin is reviewed. Emphasis is placed on (1) the calculation method used for deriving the reperfusion indices; (2) the sensitivity and the specificity of the reperfusion indices; (3) the minimum turn-around time needed to produce the reperfusion indices (depending on the practicability of the analytical and calculation methods and their applicability in an em