Elena Guerra

Biological Variation Estimates Obtained from 91 Healthy Study Participants for 9 Enzymes in Serum

BACKGROUND We sought to develop estimates of biological variation (BV) for 9 enzymes in blood serum as part of the European Biological Variation Study. METHODS Ninety-one healthy study participants (38 male and 53 female, 21-69 years old) were phlebotomized in each of 10 consecutive weeks at 6 European laboratories. The same preanalytical sample-handling protocol was followed at each center before transport to San Raffaele Hospital, Milan, Italy, for analysis. Sera were stored at -80 °C before analysis in duplicate within a single run on an ADVIA 2400 Clinical Chemistry System (Siemens Healthcare) following a protocol designed to minimize analytical imprecision. Assay traceability was established using frozen sera with target values assigned by reference methods. The results were subjected to outlier analysis before CV-ANOVA to deliver valid BV estimates. Results for 9 enzymes were subsequently partitioned for graphical display allowing visual assessment of the effects of country of origin, sex, and age on BV estimates. RESULTS We found no effect of country upon the observed variation, but overall sex-related differences were evident for alanine amino transferase (ALT), γ-glutamyl transferase (GGT), and creatine kinase (CK). The following estimates for within-subject BV (CVI) and between-subject BV (CVG), respectively, were obtained: ALT: 9.3%, 28.2%; aspartate aminotransferase: 9.5%, 20.3%; GGT: 8.9%, 41.7%; alkaline phosphatase : 5.3%, 24.9%; lactate dehydrogenase: 5.2%, 12.6%; CK: 14.5%, 31.5%; amylase: 6.8%, 30.4%; pancreatic α-amylase: 6.3%, 24.9%; and lipase (LIP): 7.7%, 23.8%. CONCLUSIONS All CVI and some CVG estimates were lower than those reported in the online BV 2014 updated database. Analytical performance specifications derived from BV can be applied internationally.

The EuBIVAS Project: Within–and Between-Subject Biological Variation Data for Serum Creatinine Using Enzymatic and Alkaline Picrate Methods and Implications for Monitoring

BACKGROUND The European Federation of Clinical Chemistry and Laboratory Medicine (EFLM) European Biological Variation Study (EuBIVAS) has been established to deliver rigorously determined biological variation (BV) indices. EuBIVAS determined BV for serum creatinine using the enzymatic and alkaline picrate measurement methods. METHOD In total, 91 healthy individuals (38 males, 53 females; age range, 21-69 years) were bled for 10 consecutive weeks at 6 European laboratories. An equivalent protocol was followed at each center. Sera were stored at -80 °C before analysis. Analyses for each patient were performed in duplicate within a single run on an ADVIA 2400 system (San Raffaele Hospital, Milan). The data were subjected to outlier and homogeneity analysis before performing CV-ANOVA to determine BV and analytical variation (CVA) estimates with confidence intervals (CI). RESULTS The within-subject BV estimates [CVI (95% CI)] were similar for enzymatic [4.4% (4.2-4.7)] and alkaline picrate [4.7% (4.4-4.9)] methods and lower than the estimate presently available online (CVI = 5.9%). No significant male/female BV differences were found. Significant differences were observed in mean creatinine values between men and women and between Turkish individuals and those of other nationalities. Between-subject BV (CVG) estimates, stratified accordingly, produced CVG values similar to historical BV data. CVA was 1.1% for the enzymatic and 4.4% for alkaline picrate methods, indicating that alkaline picrate methods fail to fulfill analytical performance specifications for imprecision (CVAPS). CONCLUSIONS The serum creatinine CVI obtained by EuBIVAS specifies a more stringent CVAPS than previously identified. The alkaline picrate method failed to meet this CVAPS, raising questions regarding its future use.

Commutable Calibrator with Value Assigned by the IFCC Reference Procedure to Harmonize Serum Lactate Dehydrogenase Activity Results Measured by 2 Different Methods

BACKGROUND The availability of commutable calibrator materials may ease considerably the task of harmonizing assay results and ensuring their traceability to reference procedures. We sought to verify the commutability of potential calibrator materials and evaluate their effectiveness in harmonizing LDH results by 2 measurement methods. METHODS We measured LDH in 109 serum samples and 31 materials, including frozen serum pools (with either normal or abnormal isoenzyme patterns), commercial stabilized materials, and the ERM-AD453/IFCC reference material. We assayed LDH activity with the IFCC reference procedure and with 2 commercial methods, 1 using the lactate-to-pyruvate (LP) reaction, and the other the pyruvate-to-lactate (PL) reaction. We selected a commutable material, with LDH value assigned by the reference procedure, as a calibrator for recalculating the results for patient sera by both LP and PL, thereby making them traceable to the IFCC reference procedure. RESULTS Original values for patient sera (n = 109) by the 2 commercial methods showed a mean (SD) PL/LP ratio of 1.97 (0.03); this ratio changed to 1.06 (0.02) after recalculation of results. Linear regression of PL vs LP recalibrated values gave y = 1.108x - 9.7. At the clinically important concentration of 250 U/L (upper reference limit), the systematic difference between methods was 6.8%, which met our proposed quality specifications for inaccuracy and total error. CONCLUSIONS By properly selecting the calibrator, the results of serum LDH measurement by 2 different methods may be harmonized and made traceable to the selected highest (reference) metrological level.

Comparative Performance Assessment of Point-of-Care Testing Devices for Measuring Glucose and Ketones at the Patient Bedside

Background: Point-of-care (POC) testing devices for monitoring glucose and ketones can play a key role in the management of dysglycemia in hospitalized diabetes patients. The accuracy of glucose devices can be influenced by biochemical changes that commonly occur in critically ill hospital patients and by the medication prescribed. Little is known about the influence of these factors on ketone POC measurements. The aim of this study was to assess the analytical performance of POC hospital whole-blood glucose and ketone meters and the extent of glucose interference factors on the design and accuracy of ketone results. Methods: StatStrip glucose/ketone, Optium FreeStyle glucose/ketone, and Accu-Chek Performa glucose were also assessed and results compared to a central laboratory reference method. The analytical evaluation was performed according to Clinical and Laboratory Standards Institute (CLSI) protocols for precision, linearity, method comparison, and interference. Results: The interferences assessed included acetoacetate, acetaminophen, ascorbic acid, galactose, maltose, uric acid, and sodium. The accuracies of both Optium ketone and glucose measurements were significantly influenced by varying levels of hematocrit and ascorbic acid. StatStrip ketone and glucose measurements were unaffected by the interferences tested with exception of ascorbic acid, which reduced the higher level ketone value. The accuracy of Accu-Chek glucose measurements was affected by hematocrit, by ascorbic acid, and significantly by galactose. The method correlation assessment indicated differences between the meters in compliance to ISO 15197 and CLSI 12-A3 performance criteria. Conclusions: Combined POC glucose/ketone methods are now available. The use of these devices in a hospital setting requires careful consideration with regard to the selection of instruments not sensitive to hematocrit variation and presence of interfering substances.

Glycated albumin: correlation to HbA1c and preliminary reference interval evaluation.

*Corresponding author: Roberto Testa, MD, Experimental Models in Clinical Pathology, INRCA-IRCCS National Institute, Via Della Montagnola 81, Ancona 60127, Italy, Phone: +39 071 800 3854, Fax: +39 071 800 3556, E-mail: r.testa@inrca.it Ferruccio Ceriotti and Elena Guerra: Service of Laboratory Medicine, Ospedale San Raffaele, Milan, Italy Anna Rita Bonfigli: Scientific Direction, Italian National Research Center on Aging (INRCA), Ancona, Italy. http://orcid.org/0000-00029619-0181 Massimo Boemi and Michela Cucchi: Metabolic Diseases and Diabetology Unit, Italian National Research Center on Aging (INRCA), Ancona, Italy Nicola Di Gaetano and Gabriele Santini: IL Diagnostic R&D Department Instrumentation Laboratory S.p.A. – A Werfen Company, Milan, Italy. http://orcid.org/0000-0002-6590-5869 (N. Di Gaetano). http://orcid.org/0000-0003-2344-3237 (G. Santini) Stefano Genovese: Department of Cardiovascular and Metabolic Diseases, IRCCS Multimedica, Sesto San Giovanni, Milan, Italy. http://orcid.org/0000-0002-6085-437X Antonio Ceriello: Centro de Investigación Biomédica en Red de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM), Barcelona, Spain; and Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Barcelona, Spain Letter to the Editor

Sequential Evaluation of Serum Urate Concentrations in AIDS Patients with Infections of the Central Nervous System

Abstract To evaluate the serum urate levels in AIDS patients with infections of the central nervous system (CNS), 46 patients who had at least two measurements of urate were included. A maximum of four measurements per patient were considered: prior to the CNS involvement (U-PRIOR), at the time of CNS involvement (U-CNS), after treatment for the CNS infection (U-AFTER), and the last measurement before death (U-LAST). Serum U-CNS levels were significantly lower than U-PRIOR values (p=0.038). U-AFTER levels were higher than U-CNS in the patients who improved (p=0.25), and lower in the patients who did not improve (p=0.026). There were no significant differences among the four diagnostic groups in U-CNS measurement (p=0.29) but they were found in U-AFTER determinations (p=0.018), probably as a result of the different response to treatment. Hypouricemia seemed to be associated with lower survival periods. We conclude that hypouricemia is common in AIDS patients with CNS infections, probably as a result of increased renal losses of urate, and that it may have prognostic significance. CNS infections are associated with significant decreases in serum urate levels in comparison with previous values. The urate concentrations seem to increase after successful treatment of the CNS infections, whereas they decrease further in patients who do not improve.

Evaluation of the trueness of serum alkaline phosphatase measurement in a group of Italian laboratories

*Corresponding author: Federica Braga, UOC Patologia Clinica, ASST Fatebenefratelli-Sacco, Via GB Grassi 74, 20157 Milano, Italy, Phone: +390239042743, Fax: +390250319835, E-mail: fedebraga83@gmail.com; and Research Centre for Metrological Traceability in Laboratory Medicine (CIRME), University of Milan, Milan, Italy Erika Frusciante, Ilenia Infusino, Elena Aloisio and Mauro Panteghini: Research Centre for Metrological Traceability in Laboratory Medicine (CIRME), University of Milan, Milan, Italy Elena Guerra and Ferruccio Ceriotti: Clinical Laboratory Service, San Raffaele Hospital, Milan, Italy Letter to the Editor

Glycation gap: An additional tool for glycometabolic monitoring

BACKGROUND The determination of glycated albumin (GA) has been suggested as an additional parameter having an independent added value respect to that of HbA1c. The determination of glycation gap (gg) has also been proposed, but few studies have addressed its potential impact in the routine evaluation of glycometabolic control. METHODS A total of 157 subjects presenting normal whole blood cell count, no hemoglobin variants, normal creatinine levels and serum protein electrophoresis patterns were studied. In a second phase, a total of 205 subjects with no restrictions as those of the first phase study, were analyzed. HbA1cwas measured by capillary electrophoresis, glycated albumin by an enzymatic method and their gg were then calculated. RESULTS The correlation between HbA1c and GA for the subjects of phase 1 was strong (r=0.8927) and significant correlation between gg and age was remarked (r=0.4486). We found 17.1% of phase 2 subjects with gg falling outside the 95% prediction intervals. Various clinical conditions seemed to affect these subjects, in our experience mostly often related to impaired renal function. CONCLUSION The glycation gap may be useful to alert clinicians about patients under unstable glycemic control or when various pre-analytical conditions my affect the reliability of the measurement of GA or HbA1c.

Evaluation of the performance of an immunoturbidimetric HbA1c reagent applied to the Siemens ADVIA 2400 automatic analyzer.

OBJECTIVES A new immunochemical reagent based on latex particles and antibodies against HbA1c (Axis-Shield), using Siemens ADVIA 2400 Instrument was evaluated. DESIGN AND METHODS Intra-assay and total imprecision, interferences studies (bilirubin ~850 μmol/L, triglycerides ~16.9 mmol/L, total protein ~140 g/L, sodium cyanate ~50 mg/dL, ascorbic acid ~50 mg/dL, urea ~24.99 mmol/L, glucose ~105.46 mmol/L, rheumatoid factor ~600 U/mL), method comparison vs Capillary Electrophoresis (Sebia), Lot to Lot reproducibility, linearity and carry over were conducted on ADVIA 2400 according to CLSI protocols. Additionally, 40 samples were measured by the two methods and also by a NGSP reference lab. RESULTS CVs % obtained by intra-assay imprecision, on 3 human HbA1c specimens at different concentrations (48, 48-64 and >64 mmol/mol) in 20 replicates, were <4%. CVs % by total imprecision, performed over 20 days with 4 calibrations on 3 human HbA1c samples (48, 48-64 and >64 mmol/mol), resulted <4%. Interferences were studied on two human samples (42-53 and>64 mmol/mol) without obtaining significant biases (<10%). Methods comparison vs Capillary Electrophoresis, performed on 120 samples ranging 23-137 mmol/mol, obtaining r = 0.974 as regression coefficient and a mean bias at decision level (48 mmol/mol) <3%. The results obtained with the NGSP samples have allowed the certification of the new reagent. CONCLUSIONS The ASD reagent met the needs of clinical laboratories, fulfilling both NGSP and IFCC requirements and it is robust to endogenous interferences.

Multicenter evaluation of an enzymatic method for glycated albumin

BACKGROUND The use of glycated albumin (GA) has been proposed as an additional glycemic control marker particularly useful in intermediate-term monitoring and in situation when HbA1c test is not reliable. METHODS We have performed the first multicenter evaluation of the analytical performance of the enzymatic method quantILab Glycated Albumin assay implemented on the most widely used clinical chemistry analyzers (i.e. Abbott Architect C8000, Beckman Coulter AU 480 and 680, Roche Cobas C6000, Siemens ADVIA 2400 and 2400 XPT). RESULTS The repeatability of the GA measurement (expressed as CV, %) implemented in the participating centers ranged between 0.9% and 1.2%. The within-laboratory CVs ranged between 1.2% and 1.6%. A good alignment between laboratories was found, with correlation coefficients from 0.996 to 0.998. Linearity was confirmed in the range from 7.6 to 84.7%. CONCLUSION The new enzymatic method for glycated albumin evaluated by our investigation is suitable for clinical use.