Carlo Franzini

Capillary zone electrophoresis of serum proteins: effects of changed analytical conditions.

Abstract Analytical conditions in a system for capillary zone electrophoresis (Beckman Paragon CZE 2000) were originally selected to allow serum protein separation into five discrete protein zones, corresponding to those of conventional clinical electrophoresis. To improve the systems performance, new analytical conditions have been made available. We compared the two sets of conditions (new = y; old = x) for possible variations of results caused by the change. One hundred thirteen serum samples, covering wide intervals of values, were assayed on two twin instruments working under the old and the new conditions; results were assessed statistically and graphically. Possible clinical significance of differences was checked by comparison with the biological variation-based quality specifications for bias. Statistically significant (y-x) differences were observed for the α1-, α2- and β-globulin zones; clinically significant differences were observed for all the zones, with the exception of the γ-globulin zone. Therefore, old/new regression equations were calculated, whose reliability was assured by the wide interval of values, by the large sample size, and by the low dispersion of single values around the mean concordance estimates. Such equations may be used to convert old into new reference values, and for the intercomparison of patient results obtained under different analytical conditions.

Recommendations for the routine use of pancreatic amylase measurement instead of total amylase for the diagnosis and monitoring of pancreatic pathology.

Abstract This document reviews the scientific evidence expected to persuade clinical laboratories to substitute pancreatic amylase measurement for total amylase in cases of suspected pancreatic pathology. A substantial evidence is now available to support such change. The measurement of pancreatic amylase in serum is: 1. more sensitive and specific for the detection of pancreatic tissue damage than that of the total enzyme activity, 2. easy and quick to perform in emergency conditions, 3. analytically precise in relation to its clinical application, 4. suitable for easy transfer and comparison of results from different care delivery units, and 5. characterized by welldefined decision limits for the diagnosis of acute pancreatitis.

Plasma or serum samples: measurements of cardiac troponin T and of other analytes compared

Abstract Conflicting data in the literature concern possible differences in the immunochemical measurement of cardiac troponins, either in plasma or in serum. In order to address this specific point, 96 serum and heparin-plasma pairs were obtained for cardiac marker measurement [cardiac troponin T (cTnT); myoglobin (Myo) and creatine kinase-MB isoenzyme (CK-MB)]; 29 additional “common” analytes were measured in 77 such samples. The cardiac markers were measured by electrochemiluminescence (Elecsys 2010, Roche); the other analytes by established automated methods (Modular, Roche). Mean plasma/serum ratios for cTnT (0.95), creatine kinase-MB (1.01) and myoglobin (0.99) were comparable with those of the 29 common analytes (interval of means 0.83–1.05). The distribution of the plasma-serum differences also showed similarities between cardiac markers and other analytes. A few outlier plasma-serum differences (3–5%) were measured for both categories of analytes. Addition of heparin to serum (51 samples) caused decreased cTnT (mean ratio 0.92). In 3 of 51 such samples the cTnT decrease was more marked, but in a second sample from the same subjects (1 week later) such a prominent, heparin-induced loss of cTnT no longer appeared. In conclusion, plasma-serum differences in immuno-reactive cTnT compare with those observed for other analytes. In occasional heparin-plasma samples immunochemical measurement of cTnT may give exceptionally low values. However, in our sample group of 96 patients (cTnT lower or higher than the cut-off in, respectively, 24 and 72 patients), no misclassification occurred if plasma instead of serum cTnT values were considered.

Ethylene glycol-stabilised haemolysates as control material in haemoglobinometry

Human blood haemolysates containing ethylene glycol (final volume fraction 0.35) were prepared and stored at -20 degrees C (in the liquid state) up to 372 days. During the whole period, the total haemoglobin concentration (assayed in the material by means of the reference HiCN method) was found to be stable; spectral analysis also failed in detecting any deterioration, Hi formation being low. Good stability was also recorded on storage at 2-4 degrees C for 15 days, but only for 1-2 days at room temperature. The stabilised haemolysate is suggested as a material for long-term control of accuracy in hemoglobinometry.

Harmonization of values for serum alkaline phosphatase catalytic activity concentration employing commutable calibration materials

BACKGROUNDnHarmonization of results allows a more effective utilization of laboratory tests; we verified the feasibility of harmonizing serum alkaline phosphatase results by two methods.nnnMETHODSnPatient sera (n=106) and candidate calibration materials (n=8) were analyzed by two methods, employing either diethanolamine (DEA) or 2-amino-2-methyl-1-propanol (AMP) as phosphate-accepting buffers. Results for patient sera by the DEA method were recalculated, with either a commutable or a non-commutable calibration material, both with values assigned by the AMP method.nnnRESULTSnAfter calibration with the commutable material, the median intermethod difference (DEA-AMP) and ratio (DEA/AMP) dropped from 195 U/l to 0 U/l and from 2.47 to 1.00, respectively. When a non-commutable material was used the former became 124 U/l and the latter 1.94. After recalibration with the commutable material, linear regression and correlation analysis of DEA vs AMP values for the set of 106 patient sera gave: intercept=0.8 U/l; slope=0.997; and nonparametric correlation coefficient r=0.9995.nnnCONCLUSIONSnHarmonization of alkaline phosphatase results by AMP and DEA methods is feasible when commutable calibration materials are used in the trueness transfer process.

Selectivity and random-access in automatic analysers

Progress in automation in clinical chemistry hasfeatured in recent years a change from rigid to very flexible instruments. The latest instruments are able to perform on each sample only those tests requested, and often do so in a random sequence. The technological aspects of this progress in instrument design are described in this paper (these include computerization ofanalytical steps, multiple wavelength technology, dry chemistry systems, new designs of centrifugal analysers and capsule chemistry). Some problems with terminology (for example selective, random and batch-selective analysers) are discussed.

Comparison of seven serum assays on four automatic analysers

Serum enzyme assays used on four different analysers (Hitachi 737, Hitachi 705, Cobas-bio and RA-2X) were compared by determining the activity of seven different enzymes (AST, ALT, LD, ALP, GGT, CK and AMS). Performance checks (quality control procedure) and replications (the study of the total analytic imprecision and of its components) were conducted and the methods were compared by linear regression analysis with statistical inference on the curves following the protocols of the National Committee for Clinical Laboratory Standards (NCCLS: PSEP- 2, PSEP-3, PSEP-4). The correlation coefficients between the methods (r = 0.991-0.999), together with the other statistical parameters, indicated that the methods are well correlated on all the instruments. The total imprecision was good for all analytes, except ALT. Among the instruments tested, the RA-2X gave more variable results, although the total imprecision was acceptable. There was no relevant carry-over effect. The evaluation of performance claims indicated that the expected error did not substantially affect the results at the level of clinical decisions.

Stability constants of haemoglobin cyanide and azide measured by two-wavelength spectrophotometric method.

A two-wavelength spectrophotometric procedure for the simultaneous determination of haemoglobin and haemoglobin cyanide (HiCN) (or of haemoglobin and haemoglobin azide (HiN3)) concentrations in mixtures has been developed and applied to the determination of the stability constants of HiCN and HiN3. The analytically reliable procedure allowed stability constants to be estimated with about 10% (relative standard deviation, coefficient of variation) uncertainty. Values of 1.9 X 10(6) and 2.0 X 10(5) 1 X mol-1 were obtained for HiCN and HiN3, respectively. These results are discussed in relation to the optimal composition of the reagents for blood haemoglobin assay by the two methods.

Diluted human serum as a suitable diluent in the preparation of bilirubin reference solutions

From a concentrated solution of bilirubin in dimethylsulfoxide (about 13 mmol/1, standardized spectrophotometrically) and a serum diluted 1/3 (whose native bilirubin is lowered by exposure to phototherapy lamps) a bilirubin-enriched serum is prepared (bilirubin concentration about 130 mumol/1 = 7.5 mg/dl) wherein all the bilrubin is bound to albumin. Such an enriched serum behaves in the diazo-reaction in the same way as a bilirubin-enriched serum prepared with undiluted serum and can, therefore, be used as a working standard in serum bilirubin assays.