Multicenter evaluation of the first automated Elecsys sFlt-1 and PlGF assays in normal pregnancies and preeclampsia.

OBJECTIVES: Performance evaluation of Elecsys sFlt-1 and PlGF assays. DESIGN AND METHODS: Within-, between-run, total imprecision, functional sensitivity, inter-laboratory comparison, method comparison and lot-to-lot reproducibility were evaluated. RESULTS: Within- and between-run CVs were below 4% for sFlt-1 >60 and PlGF > 20 pg/mL. Total imprecision CVs were below 4.3%. Functional sensitivity was < 5 pg/mL. Inter-laboratory CVs were <5%. Elecsys correlated well with Quantikine VEGF-R1 (r=0.960) and PlGF (r=0.968). Lot-to-lot comparisons yielded highly correlated results (r>0.999). In healthy pregnancies, the median levels of sFlt-1 remained constant in first (1107 pg/mL) and second trimesters (1437 pg/mL) but increased in the third trimester (2395 pg/mL), while median PlGF levels increased in the first (30 pg/mL) and second trimesters (279 pg/mL) and peaked at 29 to 32 weeks (626 pg/mL) and decreased thereafter (340 pg/mL). The sFlt-1/PlGF ratio is highest in the first trimester (median: 28) but remained constant in the second (median: 4.7) and third trimesters (median: 5.1). In PE/HELPP samples matched for gestational age the sFlt-1 levels were significantly higher (6894-34,624 pg/mL), whereas PlGF levels were lower (9.2-80 pg/mL) and the median sFlt-1/PlGF ratio is much higher (461; range: 121-2614) than in apparently healthy pregnancies (3.6; range: 0.3-105). CONCLUSION: The new Roche Elecsys sFlt-1 and PlGF immunoassay showed excellent precision and reliability. There was a clear difference in the Elecsys sFlt-1/PlGF ratio between samples obtained from women with apparently normal pregnancy at the time of blood collection and those diagnosed with PE/HELLP at the same age of gestation.

Multi-center evaluation of analytical performance of the microparticle enzyme immunoassay for sirolimus.

OBJECTIVES: This study evaluated the analytical characteristics of the new Abbott microparticle enzyme immunoassay (MEIA) for sirolimus. DESIGN AND METHODS: The protocol consisted of nine sections: evaluation of antibody specificity, linearity, detection limit, quantification limit, endogenous interferents, exogenous interferents, precision, proficiency testing panel, and method comparison. RESULTS: The mean analytical detection limit was 0.68 microg/L. The sirolimus concentration corresponding to a total CV of 20% was 1.5 microg/L. Linearity of response was demonstrated across the dynamic range of the assay. Total precision (CVs) at QC control levels from 5 to 22 microg/L ranged from 5.7 to 12.6%. Assay standardization was found to be in good agreement with LC/MS/MS as compared with target values for spiked sirolimus proficiency samples from an international sirolimus proficiency testing program. Good correlations (R values) of the immunoassay were observed in comparisons to LC/MS/MS. R values tended to be lower in comparisons with LC/UV methods. Across both LC-based methods and all study sites, there was approximately 25% overall positive slope bias due to cross reactivity of the MEIA antibody to metabolites of sirolimus. The assay cross-reactivity to metabolites of sirolimus parent drug ranged from 6 to 63%. Assay interferences were minimal with the exception of hematocrit, which presented a negative relationship to measured sirolimus concentration. CONCLUSIONS: The MEIA demonstrated acceptable analytical characteristics for use for routine monitoring of sirolimus immunosuppressive therapy, and is a viable alternative to HPLC-based methods for sirolimus monitoring.

WITHDRAWN: Multi-center evaluation of analytical performance of the microparticle enzyme immunoassay for sirolimus.

The Publisher regrets that this article is an accidental duplication of an article that has already been published in Clin. Biochem. 39 (2006) 378-386, doi:10.1016/j.clinbiochem.2006.01.017. The duplicate article has therefore been withdrawn. This article has been withdrawn consistent with Elsevier Policy on Article Withdrawal (http://www.elsevier.com/locate/withdrawalpolicy). The Publisher apologizes for any inconvenience this may cause.

Predicting the phenylalanine blood concentration from urine analyses. An approach to noninvasive monitoring of patients with phenylketonuria.

The need for regular blood-drawing in the management of chronic metabolic disorders may negatively influence the compliance of patients and their parents; noninvasive analytical procedures could well alleviate this burden. Using data obtained in six adult probands with phenylketonuria, we evaluate the feasibility of noninvasive prediction of phenylalanine blood concentrations from analysis of phenylalanine and creatinine in urine. Cross-validated regression equations correct for the significant inter-individual variation of phenylalanine fractional excretion rates. With sensitive and specific enzymatic assays for phenylalanine and creatinine, the accuracy of this noninvasive procedure may also become clinically satisfactory for the purpose of self-monitoring.

Detection limit, cut-off and specificity of an improved rapid assay for cardiac troponin T.

The performance of an improved version of the troponin T rapid test TROPT Sensitive was investigated in a multicentre evaluation at twelve centres. The detection limit and the cut-off were determined in a method comparison with Elecsys Troponin T using a total of 365 samples from patients with suspected acute coronary syndromes and 91 samples from healthy blood donors or non-cardiological patients. The analytical specificity was determined by measuring 1271 blood samples from blood donors without any myocardial injury. The test cut-off (90% of results positive) is 0.08 microg/L, and the detection limit is about 0.05 microg/L. The analytical specificity of the test is between 99.7 and 99.9%. With its small area of undefined significance between positive and negative results and its high sensitivity and specificity, TROPT Sensitive is very well suited to the reliable detection of troponin T positive patients with acute coronary syndromes.

Cardiac troponin I elevation in acute pulmonary embolism is associated with right ventricular dysfunction.

OBJECTIVES: The purpose of this study was to evaluate the prevalence and diagnostic utility of cardiac troponin I to identify patients with right ventricular (RV) dysfunction in pulmonary embolism. BACKGROUND: Right ventricular overload resulting from elevated pulmonary resistance is a common finding in major pulmonary embolism. However, biochemical markers to assess the degree of RV dysfunction have not been evaluated so far. METHODS: In this prospective, double-blind study we included 36 study patients diagnosed as having acute pulmonary embolism. RESULTS: Among the whole study population, 14 patients (39%) had positive troponin I tests. Ten of 16 patients (62.5%) with RV dilatation had increased serum troponin I levels, while only 4 of 14 patients (28.6%) with elevated troponin I values had a normal RV diameter as assessed by echocardiography, indicating that positive troponin I tests were significantly associated with RV dilatation (p = 0.009). Patients with positive troponin I tests had significantly more segmental defects in ventilation/perfusion lung scans than patients with normal serum troponin I (p = 0.0002). CONCLUSIONS: Our data demonstrate that more than one-third of patients clinically diagnosed as having pulmonary embolism presented with elevated serum troponin I concentrations. Troponin I tests helped to identify patients with RV dilatation who had significantly more segmental defects in lung scans. Thus, troponin I assays are useful to detect minor myocardial damage in pulmonary embolism.

Superiority of combined CK-MB and troponin I measurements for the early risk stratification of unselected patients presenting with acute chest pain.

BACKGROUND: Recent studies have suggested that positive troponin I tests are associated with an increased risk of cardiac death during short-term follow-up. However, it is unknown if troponin I tests alone or in addition to CK-MB measurements are superior to predict unfavorable outcome during long-term follow-up. PATIENTS AND METHODS: In a prospective, double-blind study we assessed the prevalence and prognostic value of combined troponin I and CK-MB tests in an unselected cohort of patients (n = 292) admitted to the emergency department for acute chest discomfort. Patients were grouped according to the diagnosis on discharge in those with acute myocardial infarction (1), unstable angina (2), and noncardiac chest pain (3). Six months after enrollment, death rates were obtained and follow-up interviews were performed with respect to survival, recurrence of chest pain, and myocardial infarction. RESULTS: In patients with evidence of coronary heart disease, the mortality rate for abnormal troponin I and normal CK-MB levels was 5.0%. Baseline troponin I and elevated CK-MB levels were associated with a mortality rate of 4.0%. However, the mortality rate was significantly higher (11.1%) in patients presenting with elevated troponin I and CK-MB values. In patients without myocardial infarction on admission, 10.5% with positive troponin I tests died compared to 1.6% with negative tests. The mortality rate in patients without myocardial infarction was 2.7% for patients with elevated CK-MB but normal troponin I values. In patients with both markers elevated a significantly higher mortality rate (16.7%) was found, representing a 6-fold increase in the death event rate. With the additional knowledge of troponin I values, it could be demonstrated that certain cases were misclassified as having noncardiac chest pain. At least some of the latter patients with above-normal values of troponin I were retrospectively to be reclassified as unstable angina. Acute non-Q-wave myocardial infarctions were occasionally misdiagnosed as either angina pectoris or nonischemic chest pain. CONCLUSIONS: Our data suggest the superiority of combined CK-MB and troponin I measurements in clinical practice for the early risk stratification of patients presenting with acute chest pain. In nonmyocardial infarctions, both CK-MB and troponin I convey independent prognostic information with regard to fatal outcome. Troponin I tests in addition to CK-MB measurements contribute to a lower rate of misdiagnoses.

Multicentre evaluation of the Boehringer Mannheim/Hitachi 911 Analysis System.

The analytical performance and practicability of the Boehringer Mannheim (BM)/Hitachi 911 analysis system have been assessed in a multicentre evaluation, which involved six laboratories from European countries. Analytes commonly used in classical clinical chemistry were tested in a core programme, which mainly followed the ECCLS guidelines. In addition, a satellite programme covered other analytes, such as proteins, drugs and urine analytes. In total, the study comprised more than 100 000 data items collected over a three-month period. The evaluation was supported with 'Computer Aided Evaluation' (CAEv) and telecommunications.Acceptance criteria for the results were established at the beginning of the study. Nearly all of the analytes met the imprecision limits: within-run imprecision (as CVs) was 2% for enzyme and substrate assays, 1% for ISE methods and 5% for immunoassays; between-day imprecision was 3l% for enzyme and substrate assays, 2% for ISE methods and 10% for immunoassays.No relevant drift effects (systematic deviation >/= 3%) were observed over eight hours. The methods were linear over a wide range. Sample-related and reagent-dependent carry-over can be reduced to a negligible amount by integration of a softwarecontrolled wash-step.Endogenous interferences were found for creatinine (Jaffé method) and uric acid assays (caused by bilirubin), for creatine kinase, creatine kinase MB isoform and gamma-glutamyltransferase (caused by haemoglobin), and for immunoglobulin A (caused by lipaemia)Accuracy was checked by an interlaboratory survey, recovery studies in control materials and method comparison studies. The survey showed that, with the exception of cholesterol and iron in two laboratories, the recovery of analytes did not deviate by more than 5%. Sixty-six of the 77 method comparisons performed met the acceptance criteria. The deviations of the remaining 11 results could be explained by differences in either calibration, application or by the use of different methods.Practicability was assessed using a questionnaire which covered all of the important aspects of an analysis system in the clinical laboratory. Twelve groups of attributes out of 14 were rater higher for the BM/Hitachi 911 than for the present situation in the laboratories concerned. Especially high scores were given for the versatility group.The acceptance criteria for the analytical performance of the BM/Hitachi 911 analysis system were fulfilled in all laboratory segments with few exceptions. The practicability exceeded the requirements in most of the attributes. The results of the study confirmed the usefulness of the system as a consolidated workstation in small- to medium-sized clinical laboratories and in STAT laboratories, or as an instrument for special analytes like proteins and drugs, or for urinalysis in large laboratories.

Keto acids in tissues and biological fluids: O-t-butyldimethylsilyl quinoxalinols as derivatives for sensitive gas chromatographic/mass spectrometric determination.

Quinoxalinol t-butyldimethylsilyl ethers were prepared from three branched-chain and from two aliphatic unbranched 2-keto acids. The electron impact (EI) mass spectra display pronounced [M-57]+ ions. With 39-51% of total ion current contained within them, sensitivity is greater than on chemical ionization (CI) mass spectrometry of O-trimethylsilyl derivatives. Mass spectra and chromatographic behaviour of these novel keto acid derivatives are discussed and preliminary quantitative data from rat muscle are given.

[Propionic acidemia with myelination disorders of the CNS]. / Propionazidämie mit Myelinisierungsstörungen im ZNS.

Clinical course and special diagnostic procedures in a 7 1/2 weeks old dystrophic infant with propionic acidemia are described. The disorder manifested with vomiting and diarrhea within the first week of life when the child was on a cow milk formula. Parenteral nutrition with glucose and electrolytes led to improvement. When oral nutrition with a cow milk formula was implemented again, an acute deterioration with diarrhoea and vomiting occurred. Thus, a diagnosis of cow milk allergy was suggested. There was also a severe muscular hypotony. Oral nutrition with a soybean formula did not prevent further clinical deterioration. At 7 1/2 weeks of age the patient died with symptoms of cardiogenic shock. The correct diagnosis was considered too late and confirmed post mortem. Clinical symptoms in the neonatal period like vomiting, muscular hypotony and failure to thrive should alert the physician to a possible diagnosis of a hereditary organic aciduria. Gas chromatography-mass spectrometry of urinary organics acids, in the present case, established the diagnosis. On autopsy, spongy degenerations were found in CNS.

Quantitative gas chromatographic mass spectrometric determination of mandelic acid in blood plasma. Comparison of deuterated and homologous internal standards.

Quantitative gas chromatography/mass spectrometry/selected ion monitoring of mandelic acid in plasma in the lower micromolar range has been investigated using both the deuterated compound and a homologue, 2-phenyllactic acid, as internal standards. On chromatography of the TMSi-ester ethers, the latter is less stable than the former. The chromatographic isotope effect observed for deuterated mandelic acid does not suggest a carrier function for this compound. Normal plasma levels of mandelic acid are about 0.5 microM. Only a minor portion of phenyramidol is metabolized to mandelic acid. Preliminary in vivo data indicate the presence of a stereoselective transport system for D(-)-mandelic acid in gastrointestinal tract and possibly kidney.

[Accuracy of HDL cholesterol measurements]. / Richtigkeit der HDL-Cholesterinmessung.

The widespread use of different methods for the determination of HDL-cholesterol (in Europe: sodium phosphotungstic acid/MgCl2) in connection with enzymatic procedures (in the USA: heparin/MnCl2 followed by the Liebermann-Burchard method) but common reference values makes it necessary to evaluate not only accuracy, specificity, and precision of the precipitation step but also of the subsequent cholesterol determination. A high ratio of serum vs. concentrated precipitation reagent (10:1 V/V) leads to the formation of variable amounts of delta-3.5-cholestadiene. This substance is not recognized by cholesterol oxidase but leads to an 1.6 times overestimation by the Liebermann-Burchard method. Therefore, errors in HDL-cholesterol determination should be considered and differences up to 30% may occur between HDL-cholesterol values determined by the different techniques (heparin/MnCl2 - Liebermann-Burchard and NaPW/MgCl2-CHOD-PAP).

Renal transport of aromatic acids in patients with phenylketonuria.

Renal clearance of phenylpyruvic acid is maximal at a plasma concentration of 40-60 mumol/l. This concentration is obtained with plasma phenylalanine concentrations of 1.0-1.2 mmol/l, the threshold for separating classical phenylketonuria from phenylketonuria variants.

Hydrogen exchane at the beta-carbon of amino acids during transamination.

The hydrogen exchange at the Beta-carbon of L-alanine, L-glutamate and L-asparate with water has been examined during transamination catalyzed by glutamic-oxaloacetic transaminase and by glutamic-pyruvic transaminase. A significant hydrogen exchange at the Beta-carbon has been demonstrated during incubation of L-[3-3H]alanine + glutamic-pyruvic transaminase, L-[3-3H]alanine + alpha-oxo-glutarate + glutamic-pyruvic transaminase, L-[3-3H]glutamate + glutamic-oxaloacetic transaminase, L-[3-3H]glutamate + oxaloacetate +glutamic-oxaloacetic transaminase, and L-[3-3H]glutamate + pyruvate + glutamic-pyruvic transaminase as shown by the appearance of 3H2O. No hydrogen exchange at the Beta-carbon of L-glutamate occurred during incubation of L-[3-3H]-glutamate with glutamic-pyruvic transaminase alone. The hydrogen exchaned at the Beta-carbon of L-glutamate coincides with transamination as demonstrated by nuclear magnetic resonance studies of 2H2O-L-glutamate exchange during transamination by glutamic-oxaloacetic transaminase and glutamic-pyruvic transaminase. No hydrogen exchange at the Beta-carbon occurred during transamination of L-aspartate by glutamic-oxaloacetic transaminase as shown by nuclear magnetic resonance spectroscopy and confirmed by nuclear magnetic resonance simulation studies. The results are discussed with special reference to the different equilibria between the pyridoxal form and the pyridoxamine form of glutamic-oxaloacetic transaminase and of glutamic-pyruvic transaminase.