Serum immunoreactive pancreatic lipase and cationic trypsinogen for the assessment of exocrine pancreatic function in older patients with cystic fibrosis.

Indirect and qualitative tests of pancreatic function are commonly used to screen patients with cystic fibrosis for pancreatic insufficiency. In an attempt to develop a more quantitative assessment, we compared the usefulness of measuring serum pancreatic lipase using a newly developed enzyme-linked immunosorbent immunoassay with that of cationic trypsinogen using a radioimmunoassay in the assessment of exocrine pancreatic function in patients with cystic fibrosis. Previously, we have shown neither lipase nor trypsinogen to be of use in assessing pancreatic function prior to 5 years of age because the majority of patients with cystic fibrosis in early infancy have elevated serum levels regardless of pancreatic function. Therefore, we studied 77 patients with cystic fibrosis older than 5 years of age, 41 with steatorrhea and 36 without steatorrhea. In addition, 28 of 77 patients consented to undergo a quantitative pancreatic stimulation test. There was a significant difference between the steatorrheic and nonsteatorrheic patients with the steatorrheic group having lower lipase and trypsinogen values than the nonsteatorrheic group (P less than .001). Sensitivities and specificities in detecting steatorrhea were 95% and 86%, respectively, for lipase and 93% and 92%, respectively, for trypsinogen. No correlations were found between the serum levels of lipase and trypsinogen and their respective duodenal concentrations because of abnormally high serum levels of both enzymes found in some nonsteatorrheic patients. We conclude from this study that both serum lipase and trypsinogen levels accurately detect steatorrhea in patients with cystic fibrosis who are older than 5 years but are imprecise indicators of specific pancreatic exocrine function above the level needed for normal fat absorption.(ABSTRACT TRUNCATED AT 250 WORDS)

Multicenter evaluation of a chromogenic substrate method for photometric determination of prothrombin time.

A multicenter study of a chromogenic substrate method for photometric determination of prothrombin time was conducted in order to evaluate its clinical application. Seven laboratories participated in the study using a total of 742 plasma samples from 417 patients on oral anticoagulant therapy, 261 healthy subjects and 64 patients with different diseases especially of the liver as well as 30 patients with hereditary deficiency of coagulation factors II, V, VII, X. The chromogenic PT method was compared to a standardized coagulometric PT assay which uses the same sensitive human placenta thromboplastin calibrated against international reference preparations. A high correlation of the prothrombin ratio values of the chromogenic and the coagulometric assay was obtained in 402 plasma samples (r = 0.940; y = 1.02x - 0.1). The study showed that the chromogenic PT reagent is sensitive to deficiency of the coagulation factors of the extrinsic pathway but not affected by heparin up to 1 IU/ml because of the heparin antagonist added. The precision (coefficient of variation) of the photometric method ranged between 0.6 and 3% (intraassay CV) and between 1.4 and 5.8 (interassay CV). The International Sensitivity Index (ISI) obtained for the used lot was 1.09. The therapeutical range in percentage activity for patients in a stable phase of an anticoagulant therapy was found to be from 15 to 27 percent of normal. The results of the clinical evaluation proved the good comparability of the new chromogenic PT test with coagulometric methods, its high factor sensitivity, good reproducibility and easy performance.

Standardization activities for harmonization of test results.

In the last years the search for sensitive and specific markers of renal damage and/or renal function has conducted to the development of laboratory assays for measurement of urinary proteins such as albumin, beta(2)-microglobulin, alpha(1)-microglobulin, cystatin C, etc. Furthermore, there have been new applications of already known markers based on different, reformulated methods which often rely on more advanced technologies. It is evident that such developments are connected with analytical and interpretative problems for laboratory managers and clinicians. In this situation, it is essential that international societies develop comprehensive measures for the quality management of these assays and issue uniform and carefully elaborated guidelines to ensure optimal test utilization. International activities are also directed to the development of optimized and standardized methods as well as to the production and evaluation of appropriate reference materials and, finally, to the establishment of appropriate reference ranges and cut-off values for specific analytes. The main use of reference materials is in the transfer of their accurately assigned values to the calibrators of diagnostic companies for calibration of commercially available test systems. These international standardization activities and strategies will allow a harmonized approach to disease management using a more reliable laboratory testing based on quality and value.

Evaluation and comparison of cathodic trypsin-like immunoreactivity, pancreatic lipase and pancreatic isoamylase in the diagnosis of acute pancreatitis in 849 consecutive patients with acute abdominal pain.

In 849 patients (417 men, 432 women) consecutively hospitalized with acute abdominal pain we compared the value of serum cathodic trypsin-like immunoreactivity, pancreatic lipase (EC 3.2.1.3) and pancreatic isoamylase (EC 3.2.1.1) as diagnostic tests for acute pancreatitis. The diagnoses of acute pancreatitis (in 49 patients, 5.8%) and other diseases were made without knowledge of these enzyme values. When evaluated by means of receiver operating characteristic curves no differences were found in diagnostic performance of the three enzymes. Use of combinations of different enzymes had no advantage over single enzyme determination using discrimination analysis for evaluation. The highest efficiency was for all three enzymes 0.991 (95% confidence limits: 0.983-0.995) and for all three enzymes the discrimination value giving this efficiency was several times the upper limit of reference range: 1 779 micrograms/l for cathodic trypsin-like immunoreactivity, 831 U/l for pancreatic isoamylase and 316 micrograms/l for pancreatic lipase. None of the enzymes had any prognostic value at admission in predicting a mild or severe attack of acute pancreatitis. In conclusion, no single enzyme or combination of enzymes had any diagnostic advantage for acute pancreatitis in patients with acute abdominal pain. Thus selection of one of the three enzymes as diagnostic test of acute pancreatitis is to be based on considerations such as economy, methodological simplicity, possibility of automated assay and the time-consumption at the assay.

Quality management and international standardization programs for protein immunoassays.

The huge number of new immunoassays for the measurement of specific proteins/tumor markers commercially available at present has made necessary comprehensive measures for the quality management of such laboratory tests. Very important for the comparability of test results are international standardization programs for the establishment of consensus/reference methods as well as for the development of suitable reference materials. Such programs are presently conducted by various international organizations and professional societies. Important characteristics of such reference materials are molecular composition, purity, matrix, additives, storage conditions, stability. Internationally accepted reference materials which have been prepared, calibrated and certified will contribute to the harmonization of results with different test systems, improving their quality and clinical use.

Standardization of commercial assays for serum Apo A-I and Apo B: a consensus procedure for the calibration of reference materials.

Commercial apolipoprotein A-I and B assays show a broad variation of results. This is particularly evident for the different Apo B test kits available commercially. The cause of these differences is mainly due to the inadequate international standardization of apolipoprotein assays. A common effort is at present made by commercial organizations within the International Federation of Clinical Chemistry together with the IFCC Committee on Apolipoproteins to achieve a consensus on a practical standardization procedure for apolipoprotein measurements. The aim is to calibrate all commercially available Apo A-I and Apo B test kits using frozen serum pools (at three levels) previously standardized against primary reference materials. Secondary serum reference materials (at three different concentrations) (SSRM) will be selected among those offered by industrial organizations to serve as "International Master Calibrators". These will replace the Interim Serum Reference Materials (frozen serum pools) which cannot be delivered indefinitely. The secondary serum reference materials will be used by all commercial organizations to control the validity of their own calibrations.

Proposals from IFCC Committee on Standardization of Markers of Cardiac Damage (C-SMCD): recommendations on use of biochemical markers of cardiac damage in acute coronary syndromes.

This paper presents evidence and suggestions from the IFCC C-SMCD on the use of biochemical markers for the triage diagnosis of acute coronary syndromes. There is general agreement that both 'early' and 'definitive' biochemical markers are necessary and that these assays must be available with a turnaround time of 1 h or less. Currently, myoglobin is the marker that most effectively fits the role as an 'early' marker, whereas 'definitive' markers are cardiac troponins. Since the sensitivity of the initial electrocardiogram is only 50% for detecting myocardial infarction, the use of biochemical markers may significantly contribute to the early diagnosis. New sensitive biochemical markers, particularly the cardiac troponins, are presently the best criterion to detect the presence of small myocardial cell damage. Two decision limits are probably needed for the optimum use of troponins: a low abnormal value suggesting the presence of myocardial damage and a higher value suggesting the diagnosis of myocardial infarction. Additional studies should be performed to establish limits for each commercially available assay. Finally, it is recognized that there is no need for the use of any biochemical marker when the clinical diagnosis is unequivocal, other than for diagnosing reinfarction, estimating the infarct size, and monitoring thrombolytic therapy.

Proposals from the IFCC Committee on Standardization of Markers of Cardiac Damage (C-SMCD): strategies and concepts on standardization of cardiac marker assays.

The search for sensitive and specific biochemical markers of cardiac damage has resulted in development of methods for the measurement of cardiac markers such as myoglobin, CK-MB mass and the cardiac troponins (cardiac troponin I and cardiac troponin T). There have been new clinical applications of already known markers based on improved, reformulated methods which often depend on advanced technologies. These developments are connected with analytical and interpretative challenges for the laboratory manager and for the clinician. In this situation, it is essential that international professional societies develop comprehensive and carefully elaborated guidelines for the quality management and use of these measurements and their results. Several professional associations have formed committees working on different issues regarding the measurement of biochemical markers of cardiac damage. Recognizing the huge interest in this field and substantial diagnostic relevance of these markers, the IFCC has established the Committee on "Standardization of Markers of Cardiac Damage" (C-SMCD) in 1997 inviting the already operating American and European groups to designate some of their members into the new committee. The task of the IFCC C-SMCD is to coordinate the different activities of the national groups, with preparation of international documents and recommendations under the auspices of IFCC and to initiate various standardization activities. The establishment of consensus/reference methods as well as development of primary and secondary matrix reference materials for markers of cardiac damage are extremely important in order to achieve comparability of test results, thus leading to an effective patient care.

New perspectives in diagnosis of hemostasis disorders.

The classical coagulation analyses are performed either by using manual methods or by means of various instruments with a different degree of automation. The introduction of chromogenic peptide substrates which can be split by thrombin has led to the development of photometric assays for PT and aPTT determination independent from the fibrinogen concentration and from its conversion to fibrin. After that, turbidimetric methods for the determination of fibrinogen, thrombin time and batroxobin time have been set up allowing the use of photometry for the determination of the most important hemostaseological parameters. For such purposes a new analytical system (ChromoTimeSystem, Behringwerke AG, Marburg/FRG) based on a special instrument and reagents for chromogenic and turbidimetric methods for coagulation and fibrinolysis has been developed. The ChromoTimeSystem allows to perform by photometry all important tests for coagulation and fibrinolysis. The analytical characteristics of this new system are presented.

Quality management and standardization programs in hemostaseology.

Total quality management (TQM) of laboratory services is connected with a comprehensive system of quality assurance, which integrates quality development, quality maintenance, and quality improvement. TQM concentrates not only on analytic performance and organizational issues, including specimen collection, reporting, and interpretation of results, but focuses also on the benefits to society related to the use of specific laboratory tests in prevention, early detection, and therapy monitoring, as well as on outcome measures. A prerequisite to TQM are international and national standardization programs for the establishment of optimized and standardized methods, as well as for the development and evaluation of suitable reference materials. Reference materials that have been or are being prepared, calibrated, and certified by international and national reference institutions will certainly contribute to the harmonization of results, with different test systems in hemostaseology, improving their quality and value.

[Standardization of the thromboplastin time determination]. / Standardisierung der Thromboplastinzeit-Bestimmung.

The results of our studies prove that today it is possible for manufacturers of thromboplastin preparations to follow the international recommendations for standardization of methods for prothrombin time determination. This is valid not only for the PT reagents for coagulometric techniques but also for those for photometric techniques. New manufacturing processes allow to produce sensitive thromboplastins which can be calibrated according to the recommendations of international standardization committees. Calibrated thromboplastins enable introduction of a common scale (INR) for the control of the intensity of the oral anticoagulation. The INR system improves the comparability of the therapeutical ranges recommended from different organizations and from manufacturers of thromboplastin, so that the continuity of the oral anticoagulant therapy of patients even in different countries will be decisively improved.

Present status of standardization in coagulation assays for control of oral anticoagulation.

Recently, recommendations for the production of thromboplastins have been issued by International Committees (ICSH/ICTH) to achieve a standardization of prothrombin time assays. Following such recommendations we have standardized two new PT reagents based on coagulometric and photometric methods which rely on the same sensitive human placental thromboplastin. Both PT reagents have been optimized especially for use in oral anticoagulant therapy. They offer high sensitivity and reproducibility, accurate lot standardization and therefore internationally comparable values by the expression of the ISI value of each reagent batch.

Serum levels of alpha-1 microglobulin in recipients of renal allografts.

Serum levels of alpha 1 microglobulin (s-alpha 1 m) in 92 recipients of renal transplants were elevated during pretransplant uremia (P less than 0.001), acute rejection (P less than 0.01), and cyclosporin-induced nephrotoxicity (P less than 0.01). In patients with stable renal function, those treated with cyclosporin had higher s-alpha 1 m than those receiving azathioprine: 81 +/- 4 and 64 +/- 3 mg/l (mean +/- SEM), respectively (P less than 0.05). The serum creatinine levels were 127 +/- 5 and 115 +/- 7 mumol/l (mean +/- SEM), respectively (N.S.). Two of the patients with normal serum creatinine had normal s-alpha 1m levels. There were positive linear correlations between s-alpha 1m and serum creatinine levels during stable renal function, rejection, cyclosporin-induced nephrotoxicity, and cytomegalovirus infections (r = 0.7-0.8, P less than 0.01-0.001) and between s-alpha 1m and beta 2 microglobulin (beta 2m) during the same conditions (r = 0.5-0.8, P less than 0.01-0.001). During infections, serum creatinine and beta 2m increased (P less than 0.001), but s-alpha 1m did not. S-alpha 1m values did not distinguish between rejection and cyclosporin-induced nephrotoxicity. It is concluded that s-alpha 1m might be a valuable complement to serum creatinine levels in the evaluation of renal function in renal transplant recipients.

Relevance of markers of hemostasis activation in obstetrics/gynecology and pediatrics.

Pregnancy and puerperium are considered to be hypercoagulable states with increased incidence of thromboembolic events. During normal pregnancy, changes in the hemostatic mechanism involve increased stasis and increased coagulation factors and/or decreased levels of anticoagulant proteins such as protein C and protein S as well as enhanced thrombin generation and decreased fibrinolytic activity. The physiological or pathophysiological activation of hemostasis during pregnancy results in the generation of the so-called activation markers which increase, reflecting hypercoagulability and therefore representing an imbalance in the hemostatic system. The most interesting markers of hemostasis activation and, thus, of thrombin generation are: thrombin-antithrombin III complex (TAT), antithrombin III itself, prothrombin fragment 1+2 (F 1+2), fibrin monomer (soluble fibrin) and D-Dimer (which indicates also an increased fibrinolytic activity). Together with fibrinogen levels and platelet counts, the activation markers are useful tools in different pathological situations in pregnancy to predict and monitor the severity of the condition. Recently, a higher incidence of factor V Leiden mutation has been demonstrated in selected populations in whom thrombotic events developed during pregnancy and puerperium. Therefore, the combination of APC resistance/FV Leiden mutation and pregnancy may predict a high risk for thromboembolic phenomena. In newborns, the activation markers are elevated immediately after birth and decline to near adult levels during the first 24 h of life. During infections the activation markers are increased showing the same behavior as in the mature adult system. In neonates and children, the same etiologies can be responsible for acquired and inherited pathological hypercoagulable states as in the adult.

New trends in the field of coagulation diagnosis--new possibilities to improve monitoring of antithrombotic therapy.

Two specific and sensitive enzyme immunoassays have been developed for the measurement of TAT and PTF, respectively. The TAT-ELISA uses two different antibodies binding selectively to the corresponding antigen moieties of TAT; anti-PTF antibodies were obtained from rabbits using a synthetic peptide from the COOH-terminus of PTF. Concentration in plasma samples of healthy individuals was found to be 1.45 +/- 0.4 micrograms/l for TAT, and 0.65 +/- 0.2 nMol/l for PTF. Patients with coagulation disorders showed markedly increased concentrations of both TAT and PTF. It can be assumed that these parameters might be suitable indicators for monitoring of both anticoagulant and thrombolytic therapy.

The ontogeny of serum immunoreactive pancreatic lipase and cationic trypsinogen in the premature human infant.

We evaluated the development of the exocrine pancreas in 16 healthy preterm infants (29.3 +/- 1.6 weeks). The infants were fed breast milk with formula supplements (n = 8) or formula alone (n = 8). Growth was monitored weekly for 12 weeks then at 3, 6, 9, 12 months. At the same intervals sera were determined for pancreatic lipase and cationic trypsinogen. In addition, cord blood samples were analysed from another 33 preterm (27.6 +/- 5.2 weeks) and 75 healthy full-term infants. Serum pancreatic lipase in the cord blood of term (3.7 +/- 0.4 micrograms/l) and preterm infants (1.8 +/- 0.2 micrograms/l) was significantly below values reported for older children (10.5 +/- 0.9 micrograms/l; p less than 0.001). In the preterm infant, serum lipase was also significantly lower than values obtained at term (p less than 0.001). At birth, serum trypsinogen for preterm (16.8 +/- 1.3 micrograms/l) and term infants (23.3 +/- 1.9 micrograms/l) were below those for older children (31.4 +/- 3.7 micrograms/l; p less than 0.05). Over the first 3 weeks of life, serum lipase and trypsinogen increased significantly. From 3 weeks to 12 months of age, serum trypsinogen values remained unchanged, but serum lipase increased dramatically after 10 weeks of age. Thus, at 6 and 12 months of age, the preterm infants had significantly higher serum lipase values than infants of the same age born at term. These two pancreatic enzymes appear to show independent age-related maturation in infants born before term. The rate of maturation of lipase appears to be accelerated by exposure to the extrauterine environment.