Prenatal diagnosis of OEIS (omphalocele, bladder exstrophy, imperforate anus, clubfeet) variant associated with increased nuchal translucency and OEIS complex with ambiguous genitalia associated with corrected transposition of the great arteries: case series and review of the literature.

INTRODUCTION: The OEIS complex refers to a combination of defects consisting in omphalocele, bladder exstrophy, imperforate anus and spinal defects and represents a rare nosologic entity (from 1:200,000 to 1:400,000 pregnancies). The defect probably occurs in early blastogenesis or in mesodermal migration during the primitive streak period. MATERIALS AND METHODS: Two cases of OEIS complex diagnosed prenatally by ultrasound are reported. The medical record regarding differential diagnosis, associated anomalies, treatment and prognosis has also been sought and reported. CONCLUSION: Differential diagnosis with exstrophy-epispadias complex and/or cloacalexstrophy complex may be difficult antenatally by means of ultrasound. However, color Doppler has been proved to aid the diagnosis of bladder exstrophy by depicting the urine flow in direct communication with the abdominal cavity and has been useful in showing the course of the perivesical umbilical arteries. Prenatal 3D ultrasound with tomographic ultrasound imaging (TUI) and antenatal MR imaging might be useful adjuncts to conventional 2D scan in aiding the prenatal diagnosis of such malformation.

Results from a multicenter evaluation of the 4th generation Elecsys Troponin T assay.

BACKGROUND AND OBJECTIVE: Discrepancies between serum and heparin plasma samples have been described for many commercial troponin assays including the cardiac troponin T (cTnT) assay. Using the current 3rd generation Elecsys Troponin T immunoassay, heparin plasma cannot be recommended for the determination of cTnT due to systematic lower test results caused by a direct interference of the immunoassay by heparin. The purpose of the multicenter study was to evaluate the analytical performance of an improved 4th generation Elecsys Troponin T immunoassay with a special focus on the comparability of cTnT results determined in heparin plasma and serum. METHODS AND RESULTS: The multicenter evaluation was performed in 10 clinical laboratories according to a standardized protocol (Roche Diagnostics, Penzberg, Germany, Study No. B05P008). The Elecsys Troponin T immunoassay was performed on the Modular Analytics E170 and Elecsys 2010 systems. Intraassay imprecision (n = 21) and total imprecision (2 runs/d, 10 days, triplicate measurements) were evaluated using 2 commercial controls (Roche Diagnostics) and 6 different serum pools (cTnT: 0.0140 - 4.102 microg/L). Intraassay CVs ranged from 0.73 to 3.22%. Total imprecision CVs ranged from 3.61 to 35.45% (cTnT < 0.1 microg/L) and 1.82 to 9.09% (cTnT > 0.1 microg/L), respectively. The cut-off for myocardial necrosis was determined to be 0.03 microg/L using the 10% total imprecision CV criteria. Linearity was assessed by serial dilutions of 6 different serum samples using cTnT negative serum pools. Linearity was proven up to 21.3 microg/L (recoveries: 90% - 110%). Regression data of all comparison studies were calculated according to the method of Passing and Bablok. The method comparison between the 4th generation and the commercially available cTnT immunoassay showed highly similar results across the whole measuring range (0.01 - 25.0 microg/L): y = 1.024x -0.001, r = 0.998; n = 988. Using the commercially available cTnT reagent, the serum to heparin plasma comparison yielded a systematic bias to approximately 8% lower cTnT results in heparin plasma. However, suitable comparability was obtained using the 4th generation Elecsys cTnT assay. The regression analysis (serum vs. heparin plasma) across the studied measuring range (cTnT: 0.01 - 14 microg/L) yielded the following equation: y = 0.975x + 0.001; r = 0.986; n = 403. However, rare individual serum to matched heparin plasma samples still yielded poor comparability (deviation > 20%) using the 4th generation Elecsys Troponin T immunoassay. CONCLUSION: Our data confirm an excellent analytical performance of the improved troponin T immunoassay. Most importantly, no systematic bias between cTnT results determined in serum and heparin plasma was observed from data obtained in 7 evaluation sites. The performance of the 4th generation Elecsys Troponin T assay is therefore comparable to other commercially available troponin immunoassays. Further studies are necessary to investigate the cause of poor comparability of cTnT results in rare individual serum to matched heparin plasma samples.

The exciting story of cardiac biomarkers: from retrospective detection to gold diagnostic standard for acute myocardial infarction and more.

This paper reviews the history of the contribution of the laboratory medicine to clinical cardiology and discusses the most important steps in this field. Until 20 years ago, the clinical laboratory only placed at the cardiologist's disposal a few assays for the retrospective detection of cardiac tissue necrosis, such as enzymatic methods for creatine kinase and lactate dehydrogenase activities. However, in the latter part of the 20th century, highly sensitive and specific assays, such as cardiac troponins, as well as assays for markers of myocardial function, such as cardiac natriuretic peptides, rapidly changed the scenario of clinical management of patients with cardiac diseases, assigning to the laboratory a pivotal role in the overall diagnostic flow. This is witnessed by the recent incorporation of these markers into international guidelines and in the redefinition of myocardial infarction. For the foreseeable future, new serum markers of myocardial ischemic, i.e. reversible, injury or related to coronary plaque instability and disruption are expected.

Standardization in laboratory medicine: new challenges.

The primary goal of Laboratory Medicine is to provide information that is useful to assist medical decision-making and permits optimal health care. This type of information should be independently obtained of the measurement test kits and instruments, and also of the laboratory where the procedure is carried out. It is therefore important to achieve a level of comparability of laboratory results among the many measurement procedures available so that results are harmonized and interchangeable over space and time. The standardization of measurements is therefore of high priority. In recent years, numerous efforts have been made at the international level under the auspices of the IFCC and other organizations to standardize measurement results for many important analytes, e.g. enzymes, cardiac proteins, etc. The aim of this review is to discuss some concepts related to the achievement of standardization by the implementation of a metrologically correct measurement system, providing some examples on how these concepts can be applied in Laboratory Medicine.

Serum isoforms of creatine kinase isoenzymes.

The CK-2 and CK-3 isoenzymes of human serum creatine kinase (CK) can be further subdivided into five isoforms (subforms derived from the same isoenzyme). Three are derived from CK-3 and two from CK-2. The formation of these isoforms is a postsynthetic phenomenon brought about by a serum carboxypeptidase that acts on the M monomer of the enzyme. Sera from healthy subjects contain CK-3(1) as the dominant isoform with lesser amounts of CK-3(2) and CK-3(3). Following damage of muscle tissue, the serum isoform distribution changes as a result of the increased release of CK enzyme. This provides more diagnostic information concerning acute myocardial infarction and other muscle diseases than is available from routine CK isoenzyme analysis.

Aspartate aminotransferase isoenzymes.

Aspartate aminotransferase (AST, EC 2.6.1.1) exists in human tissues as two distinct isoenzymes, one located in the cytoplasm (c-AST), and the other in mitochondria (m-AST). Striated muscle, myocardium, and liver tissues are the main sources of AST. A growing body of information suggests that determination of AST isoenzymes in human serum is useful in evaluating damage to some of these organs. In hepatic disease, the test is used to assess liver necrosis and for determining prognosis. It may also assist in identifying patients with active alcoholic liver disease. In patients with acute myocardial infarction, measurement of AST isoenzymes provides diagnostic information that differs from that obtained by determination of total creatine kinase and lactate dehydrogenase enzymes, and their isoenzymes.

Present issues in the determination of troponins and other markers of cardiac damage.

OBJECTIVE: To review some of the recently proposed improvements and the corresponding apparent issues in the field of biochemical markers of cardiac damage. CONCLUSIONS: The continuous development of new analytical tools for the biochemical evaluation of patients with suspected myocardial injury brings without doubt new challenges of careful technological evaluation, implementation, and standardization but it may also provide a unique opportunity to markedly enhance our diagnostic performance in the clinical setting of acute coronary syndrome.

Time course of changes in serum activity of the P3 isoform of pancreatic amylase isoenzyme in patients with acute pancreatitis.

We studied the temporal sequence of serum P3 isoamylase changes in patients with acute pancreatitis, in comparison with serum pancreatic lipase (LPS) activity. Twenty-four hospitalized patients with acute pancreatitis, the onset not more than 12 h previously, were admitted to the study. Serum samples were collected daily for 6 days after onset for enzyme determinations. Measurements in the 48-h period after the onset of acute pancreatitis showed clinical sensitivities of total amylase, P3 isoform and LPS of 100%. During the following period, LPS and P3 isoform activities decreased with time with parallel changes, still showing diagnostic sensitivity significantly higher than total amylase. P3 isoform and LPS appear to be interchangeable markers of pathological release of pancreatic enzymes into the bloodstream during acute pancreatitis. However, LPS determination is more convenient because of its simplicity and shorter turnaround time.

Significance of various parameters derived from biological variability for lipid and lipoprotein analyses.

Analytical, within-subject, and between-subject components of variability have been determined for total cholesterol, LDL-cholesterol, HDL-cholesterol, and its HDL2 and HDL3 subfractions, triglycerides, apolipoproteins A-I and B, and lipoprotein(a) in serum specimens from a cohort of 10 healthy subjects over a 1-month period. From these data, we have calculated the desirable analytical imprecisions, the indices of individuality, the critical differences for significant change detection, and the number of specimens required to estimate the homeostatic set-point of an individual. Practically, the analytical goal for imprecision was not achieved for HDL2 subfraction, lipoprotein(a), and direct LDL-cholesterol determination (obtained vs. theoretical analytical CV, 8.2% vs. 5.9%, 7.4% vs. 3.8%, and 3.2% vs. 2.6%, respectively). All analytes had marked individuality, showing that the use of population-based reference values is inadequate for their interpretation. The applicable differences required for two results to be significantly different (p < or = 0.05) are total cholesterol: 10%; triglycerides: 49%; LDL-cholesterol: 16%; HDL-cholesterol: 14%; HDL2-cholesterol: 40%; HDL3-cholesterol: 16%; apolipoprotein A-I: 11%; apolipoprotein B: 12%; and lipoprotein(a): 29%. Furthermore, it should be clear that population screening for the assessment of risk of coronary artery disease by means of some of the assays studied would result in a significant number of patients requiring analysis of multiple specimens.

Evaluation of a new method for cholinesterase determination.

We report the evaluation of a new commercially available assay system for determination of the catalytic activity of serum cholinesterase (EC 3.1.1.8) and application of the method to a centrifugal fast analyzer. Serum cholinesterase activity is determined at 30 degrees C using para-hydroxybenzoylcholine as substrate. This reaction is coupled to a second reaction using para-hydroxybenzoate hydroxylase (EC 1.14.13.2) as coupling enzyme. Enzyme activity is measured kinetically by monitoring the decrease in absorbance at 340 nm of NADPH in the second reaction. The procedure is precise and the results obtained from normal and pathological sera show good correlation with those obtained by the alternative procedures employing propionylthiocholine, butyrylthiocholine and benzoylcholine as substrates. The reference range for 700 healthy subjects was estimated to be 140-345 U/L (95% central range, determined non-parametrically), with significant difference between males and females (155-353 U/L for men and 134-323 U/L for women, p less than 0.001).

An immunochemical procedure for determination of creatine kinase 3(1) (serum-specific) isoform in human serum evaluated.

Changes in the proportions of individual isoforms of creatine kinase (CK) in serum promptly reflect both myocardial infarction and coronary reperfusion. A new commercial kit has been introduced for measuring CK-3(1) isoform in serum (ISOFOR-MM, International Immunoassay Labs.). This is an immunochemical assay containing CK-3(1) specific monoclonal antibody, bound to magnetizable particles, used to immunoextract this isoform. The CK activity of the sample is measured before and after immunoextraction and the difference in the two values gives the measure of CK-3(1). Extraction of CK-3(1) was complete at less than or equal to 1200 U/L. Analysis of between-day imprecision gave CV between 2.9-7.9%. The method was not susceptible to interference by CK-3(2) and CK-3(3) isoforms, CK-2 isoenzyme, or mitochondrial CK. Reference interval for CK-3(1) (expressed as percent of total CK-3) was 42-69%. Correlation between percent CK-3(1) by isoform electrophoresis (x) and evaluated procedure (y) was y = 0.83x + 7.6, with r = 0.957 (n = 40). The ISOFOR-MM performed well enough in this evaluation to replace electrophoresis or isoelectric focusing for measurement of CK-3(1) isoform.

Evaluation of the direct potentiometric method for serum chloride determination--comparison with the most commonly employed methodologies.

A new instrument (NOVA 4 + 4) designed to measure serum chloride by ion-selective electrode (ISE)-direct potentiometry (without sample dilution) was evaluated and compared with two widely used methodologies for chloride determination. Mean values obtained with the Nova were significantly higher than those obtained by a colorimetric method and by coulometric titration. Unlike these procedures, which are performed after sample dilution, direct potentiometry was unaffected by changes in plasma water caused by hyperlipemia and/or high protein concentrations. The only important interference was falsely high choride caused by administered bromide or iodide. Therefore, the direct potentiometric method should be more accurate than the colorimetric or coulometric procedures, especially when plasma lipids or proteins are very high.

Nonenzymic glycation of apolipoprotein B in patients with insulin- and noninsulin-dependent diabetes mellitus.

OBJECTIVE: To evaluate the level of nonenzymatic glycation of apolipoprotein B in patients with insulin and noninsulin dependent diabetes mellitus. METHODS: Using a method based on a combination of affinity chromatography and immunonephelometry, we measured the concentration of glycated apolipoprotein B (apo B) in serum of 140 diabetic patients, 43 insulin-dependent (IDDM), and 97 noninsulin-dependent (NIDDM), and 45 nondiabetic control subjects. RESULTS: Although total apo B concentration in serum was significantly increased only in NIDDM patients, both groups of diabetics showed higher percentages of glycated apo B (IDDM, 4.84 +/- 0.8%; NIDDM, 5.61 +/- 1.1%) than did control subjects (4.28 +/- 1.0%), the greatest percentage (5.80%) being found in patients with diabetic nephropathy. No significant correlations were found between glycated apo B and the traditional parameters of glycemic control, such as glycated hemoglobin and fructosamines, and direct influence by sudden plasma glucose fluctuations on apo B glycation was not shown either, perhaps for a low inherent glycability of this apolipoprotein. CONCLUSIONS: It is unclear if these low proportions of glycated apo B in vivo may significantly affect lipoprotein metabolism assuming a pathophysiological role in atherogenesis.

Diagnostic value of four assays for lipase determination in serum: a comparative reevaluation.

We assessed the diagnostic value of four commercially available methods for determining pancreatic lipase (LPS) in serum (the turbidimetric procedure from Boehringer, two enzymatic approaches from Kodak and Poli, and an immunochemical assay) in a population of 46 hospitalized patients with acute abdominal pain. In 31 cases (67.4%), the final diagnosis was acute pancreatitis. When evaluated by means of receiver-operating characteristic (ROC) curves, no significant differences were found among the procedures. Concerning clinical efficiency, all the assays had values equal to or greater than 90%. Using the calculation of the overlap index (OI) as a statistical approach to quantify the clinical utility of various LPS assays, the test having the greatest potential for differentiating between patients with and without acute pancreatitis was the turbidimetric assay (OI = 0.14).

Cardiac troponin elevations in chronic renal failure: prevalence and clinical significance.

OBJECTIVES: The aim of the study was investigate the prevalence of abnormal values of cardiac troponin T (cTnT) and cardiac troponin I (cTnI) in patients with chronic renal failure (CRF) and their clinical significance. DESIGN AND METHODS: We investigated the concentrations of cTnT and cTnI in 49 CRF patients without heart disease or diabetes. Cardiac TnT values were measured with a second generation immunoassay and cTnI with two immunoassays with different analytical sensitivity. All CRF patients underwent regular clinical follow-up over a 18-month period. RESULTS: No patients with CRF had elevated values of cTnI when measured with one assay and only 2 patients displayed minimally elevated values with the second assay. In contrast, 23 CRF patients (47%) displayed cTnT concentrations elevated above the upper reference limit. The elevated cTnT values observed were below the values detected in acute myocardial infarction and were not associated with adverse cardiac events during follow-up. CONCLUSIONS: Mildly elevated cTnT concentrations are common in patients with CRF and do not appear to be associated with adverse coronary events.

Serum enzymes in acute myocardial infarction after intracoronary thrombolysis.

Serum kinetics of total creatine kinase (CK), CK-MB isoenzyme, aspartate aminotransferase (AST), lactate dehydrogenase (LD) and alpha-hydroxybutyrate dehydrogenase (HBD) activities were studied in twenty patients with acute myocardial infarction randomly assigned to receive either intracoronary urokinase (group A) or conventional (control) therapy (group B). The temporal characteristics of enzyme changes described were the time lag from onset of chest pain until maximum catalytic concentration value, the rate at which enzymes are released into blood, the peak value of the serum enzyme curves and (d) the fractional disappearance rate (Kd) for each enzyme considered. Thrombolytic treatment induced earlier peak times in group A: for CK, 10.8 vs 27.0 h, for CK-MB, 10.4 vs 23.1, for AST, 13.9 vs 31.3, for LD, 24.4 vs 49.1, and for HBD, 20.5 vs 48.5 (for all enzymes, p less than 0.001). The maximal rate of release for the enzymes was at least twofold greater in group A. Enzyme peak activities and Kd were not significantly different between the groups. The most significant discrimination between the two groups was obtained with AST peak time (Hartz overlap index (Oi) = 0.11) and CK-MB peak time (Oi = 0.12).

Diagnostic application of CK-MB mass determination.

Recent advances in analytic techniques have increased the diagnostic value of creatine kinase MB (CK-MB), enabling earlier and more sensitive results. The CK-MB mass immunoassays, that utilise the monoclonal anti-CK-MB in conjunction with anti-M or anti-B antibodies, are able to measure accurately small changes during the early hours after myocardial infarction (MI). CK-MB has two main limitations in diagnosing MI neither of which however undermines its established clinical value: CK-MB is not perfectly specific to cardiac injury, with increase occurring also during massive musculoskeletal injury; furthermore, the early release pattern of CK-MB limits its value for the late MI diagnosis. For the foreseeable future evidence is compelling for greater access to rapid testing capabilities in emergency situations, using protocols incorporating CK-MB mass evaluation together with other biochemical markers, i.e. myoglobin and troponins.

Recent approaches in standardization of cardiac markers.

The development of commercial assays for the determination of new cardiac proteins has been one of the most important innovations in the field of cardiovascular diagnostics in the last decade. This significant and sudden advancement has however led to some analytical and interpretative problems. There are problems in test standardization, imprecision, interference and preanalytical variability. We also need to standardize utilization of biomarkers in diagnosis and management of acute cardiac syndromes and clearly define decision thresholds. Powerful tests, such as cardiac markers, on which critical decisions will rest, need highly reliable methods. The feeling is that some assays are inadequately appraised prior to their introduction in clinical use. More studies are needed to implement new devices in the laboratory routine, and only well documented assays should be used in hospital-based laboratories. The technology to address many analytic problems is at hand, but commitment on the part of manufacturers and their customers in the laboratory community is essential.

Determination of aspartate aminotransferase isoenzymes in hepatic diseases--preliminary findings.

The study sought to establish a relationship between the AST isoenzyme levels in serum and degree of hepatic damage, by using a new and simple immunochemical method for the differential determination of the isoenzymes. Sixty-nine patients with various hepatic diseases were studied. During hepatic damage, cytoplasmic isoenzyme (s-AST) is found in greater quantities than mitochondrial isoenzyme (m-AST), but the m-AST level increases to a greater extent in acute liver diseases. However, m-AST in alcoholic hepatitis is higher than expected from the total AST (t-AST) values. The ratio of m-AST to t-AST seems to discriminate alcoholic hepatitis from other liver diseases.