C. Prontera

CLERICO, ALDO

EMDIN, MICHELE

G. C. Zucchelli

L. Zyw

PASSINO, Claudio

S. Storti

Archivio della ricerca della Scuola Superiore Sant'Anna

during the same time frame is beyond the scope of thisobservational study.Our study highlights the fact that isoprostane concen-trations have the same limitations as other measures ofobstetric outcome, such as pH and Apgar score: i.e., theyare increased in neonates with HIE, but may also beincreased in apparently healthy infants and may bewithin reference values in infants who die from acutecauses or in whom a pathology manifests after delivery.We are grateful to all medical and nursing staffs in thelabor wards and neonatal units at Prince of Wales Hospi-tal, The First Affiliation Hospital of Medical College ofJinan University, and The Women’s & Children’s Hospitalof Guangdong Province for case recruitment and samplecollection. The study was supported by the Hong KongSpecial Administration Region Research Grant Council(CUHK4329/99M). All contributors and guarantors inthis study are independent from the funding agency, andthe views expressed here are those of the authors. Thisreport was partially awarded (Asian-Pacific Congress ofClinical Biochemistry Regional Service Award) and rep-resented at the 10th Asian-Pacific Congress of ClinicalBiochemistry and 42nd Annual Scientific Conference ofthe Australasian Association of Clinical Biochemists inPerth, Australia, during September 2004.References1. Ferguson SD, Gehrke T. Deaths related to intrapartum asphyxia. Audit in oneunit found neonatal care to be suboptimal at weekends [Letter]. BMJ1998;316:1318–9.2. Macintosh M, Lee C. Deaths related to intrapartum asphyxia. Intrapartumdeath rates in England 1993–5 did not show consistent peaks or troughs[Letter]. BMJ 1998;316:1319.3. Murphy DJ. Deaths related to intrapartum asphyxia. Denominators areneeded before conclusions can be drawn [Letter]. BMJ 1998;316:1319.4. Luckas M, Thomson A, Aird I. Deaths related to intrapartum asphyxia.Consultant expansion in obstetrics and gynaecology is not fast enough[Letter]. BMJ 1998;316:1319.5. Bastian H, Keirse MJNC, Lancaster PAL. Perinatal death associated withplanned home birth in Australia: population based study. BMJ 1998;317:384–8.6. Barela TD, Johnson JD, Hayek A. Metabolic acidosis in the newborn period.Clin Endocrinol Metab 1983;12:429–46.7. Low JA, Panagiotopoulos C, Derrick EJ. Newborn complications after intra-partum asphyxia with metabolic acidosis in the term fetus. Am J ObstetGynecol 1994;170:1081–7.8. Socol ML, Garcia PM, Riter S. Depressed Apgar scores, acid-base status,and neurologic outcome. Am J Obstet Gynecol 1994;170:991–9.9. Rogers MS, Wang W, Mongelli M, Pang CP, Duley JA, Chang AMZ. Lipidperoxidation in cord blood at birth: a marker of fetal hypoxia during labour.Gynecol Obstet Invest 1997;44:229–33.10. Wang CC, Rogers MS. Lipid peroxides in cord blood: the effects of umbilicalnuchal cord. Br J Obstet Gynaecol 1997;104:251–5.11. Liu BY, Wang CC, Lau TK, Chu CY, Pang CP, Rogers MS, et al. Meconiumstained liquor (MSL) during labor is associated with raised neonatal cordblood 8-iso-prostaglandin F2 concentration. Am J Obstet Gynecol 2004;192:289–94.12. Mongelli JM, Wang CC, Wang W, Pang CCP, Rogers MS. Oxygen free radicalactivity in the second stage of labor. Acta Obstet Gynecol Scand 1997;76:765–8.13. Rogers MS, Mongelli JM, Tsang KH, Wang CC, Law KP. Lipid peroxidation incord blood at birth: the effect of labour. Br J Obstet Gynaecol 1998;105:739–44.14. Wang W, Pang CC, Rogers MS, Chang AMZ. Lipid peroxidation in cord bloodat birth. Am J Obstet Gynecol 1996;174:62–5.15. Hoffman SW, Rzigalinski BA, Willoughby KA, Ellis EF. Astrocytes generateisoprostanes in response to trauma or oxygen radicals. J Neurotrauma2000;17:415–20.16. Rogers MS, Chang AMZ. Perinatal asphyxia: a Bayesian analysis of predic-tion and prevention. J Obstet Gynaecol 1991;11:34–40.17. Rogers MS, Murray HG, Wang CC, Pennell CE, Turner A, Yan P, et al.Oxidative stress in the fetal lamb brain following intermittent umbilical cordocclusions: a path analysis. Br J Obstet Gynaecol 2001;108:1283–90.DOI: 10.1373/clinchem.2004.047241Comparison of a Fully Automated Immunoassay with aPoint-of-Care Testing Method for B-Type NatriureticPeptide, Concetta Prontera, Simona Storti, Michele Emdin,Claudio Passino, Luc Zyw, Gian Carlo Zucchelli, and AldoClerico* (Consiglio Nazionale delle Ricerche Institute ofClinical Physiology, Pisa, Italy; * address correspondenceto this author at: Laboratory of Cardiovascular Endocri-nology and Cellular Biology, CNR Institute of ClinicalPhysiology, Via Trieste 41, 56125 Pisa, Italy; fax 39-0585-493601, e-mail clerico@ifc.cnr.it)The clinical relevance of B-type natriuretic peptide (BNP)as a diagnostic tool and prognostic marker in patientswith cardiovascular diseases has been confirmed recently(1–3). Over the last 5 years, several immunoassay meth-ods for the measurement of BNP, which share someanalytical characteristics, such as full automation, turn-around time 60 min, lower imprecision, and/or betteranalytical and functional sensitivity, have become com-mercially available (1, 4). The analytical performance anddiagnostic accuracy of immunoassays for BNP were com-pared recently (5, 6). In the present study, we evaluatedthe analytical performance and diagnostic accuracy of anautomated immunochemiluminescent assay for BNP(ACCESS System; Beckman Coulter). We also comparedits performance and accuracy with the TRIAGE BNP test(Biosite). The 2 immunoassays use as antigen the intactBNP 1–32 peptide and the same mouse anti-human BNPantibodies. The capture antibody (Scios) recognizes thepeptide ring, whereas the detection antibody (Biosite)recognizes an epitope between amino acids 5 and 10 at theNH2 terminus (6, 7). All statistical analyses were per-formed by use of parametric tests after log transformationof the original data. Nonparametric tests were also per-formed and showed identical statistical trends.We collected blood samples from 91 healthy individu-als [51 women and 40 men; mean (SD) age, 43.2 (13.4)years; range, 16–71 years] and 214 patients with idio-pathic or secondary cardiomyopathy (only 21 with a leftventricular ejection fraction 50%). Blood was collectedbetween 0800 and 0900 after the individuals had fastedovernight and had rested for 20 min in a supine position.Immediately after withdrawal, blood samples (8–10 mL)were placed in ice-chilled disposable polypropylene tubescontaining EDTA (1 mg/mL of plasma). Plasma sampleswere obtained shortly after venipuncture by centrifuga-tion for 15 min at 4 °C and then were frozen and stored at20 °C in 0.5-mL aliquots until being assayed.We found that plasma samples can be kept at 37 °C for1274 Technical Briefs4 h without a significant decrease in measured (on anACCESS system) BNP concentration [mean (SD) mea-sured value, 97.8 (5.3)% of the original concentration;mean difference from the original concentration, 10.3 ng/L;95% confidence interval (CI), 20.4 to 41.1 ng/L (P 0.1147); median measured value, 790 ng/L (range, 37–2904ng/L); n  15]. We observed a slight but significant differ-ence when plasma samples were kept at 4 °C for 24 h [meanmeasured value 89.0 (5.0)% of the original concentration;mean difference from the original concentration, 27.5 ng/L;95% CI, 13.7–41.4 ng/L (P 0.0001); median measuredvalue, 160.5 ng/L (range, 12–824 ng/L); n  24].We assessed the detection limit of the ACCESS byrepeatedly measuring (n  20) the calibrator with a BNPconcentration of 0 ng/L in 3 different runs. The mean (SD)measured concentration was 0.3 (0.1) ng/L (range, 0.3–0.4ng/L). We confirmed the results of a previous study fromour laboratory indicating that the detection limit of theTRIAGE is 8 ng/L (manufacturer-reported detectionlimit, 5 ng/L) (8 ). The imprecision profiles of the TRIAGEand ACCESS, obtained by repeated measurements over20 different days of several plasma samples (7 on theTRIAGE and 12 on the ACCESS) with BNP concentrationscovering the analytical working range, are shown in Fig.1A. According to the IFCC goals for desirable (15% totalCV) and optimum (10% total CV) imprecision (7 ), the 2methods had similar functional sensitivities (10 ng/Land 30 ng/L, respectively). The imprecision for sampleswith BNP values 10 ng/L was not satisfactory in eithermethod. We assessed the within-run and total imprecisionof the ACCESS according to the NCCLS EP5-A guideline(including experimental protocol, number of measure-ments, and data calculation) (9 ) by repeatedly assaying 2plasma samples with BNP concentrations of 52.6 and 1095ng/L in duplicate on 20 different working days (both inthe morning and in the afternoon). For the 2 samples,within-run imprecision was 3.4% and 1.6%, respectively,and total imprecision was 8.4% and 5.9%.After exclusion by statistical analysis of the 17 sampleswith BNP values 5 ng/L in the TRIAGE assay, we founda close linear relationship (R  0.919; n  287) between the2 methods: ACCESS 62.5 (14.1) 0.932 (0.024) TRIAGE (P0.0001 for both). The Bland–Altman plot of the same dataconfirmed that there was a difference between the valuesmeasured with the 2 immunoassays; this difference in-creased proportionally as the measured concentration in-creased (Fig. 1B) (10). There was a slight but significantmean difference between the values [mean (SD) differencefor TRIAGE  ACCESS, 45.2 (217.8) ng/L (minimum–maximum, 1152 to 1667 ng/L; P 0.0001); mean %difference,0.2% (minimum–maximum,2.0% to 1.7%)].All 91 healthy persons were nonobese, normotensive,and free from acute or chronic diseases. Moreover, all hadvalues within the reference intervals for creatinine, urea,glucose, uric acid, albumin, electrolytes, cardiac markers,and hemoglobin as well as for erythrocyte and leukocyteFig. 1. Total imprecision profiles (A) and Bland–Altman difference plot(B) for the tested immunoassays.(A), the CV values corresponding to desirable imprecision (15% CV) and optimumimprecision (10% CV), respectively, are indicated with dashed lines. (B), thedifference plot shows the difference between methods as a percentage (y axis)plotted vs the mean value (x axis). Solid horizontal lines indicate the meandifference and the 	 3 SD values. The dashed line indicates no difference.Table 1. Characteristics of the heart failure patientsenrolled for the evaluation of diagnostic accuracy.AllNYHA classI-II III-IVNo. of patients 193 122 71Mean (SD) age, years 64 (13) 62 (13) 67 (12)F/M, n 45/148 28/94 16/55Mean (SD) BMI,a kg/m2 26 (4) 27 (2) 25 (3)Idiopathic/postischemic/secondary cardiomyopathy,%49/36/15 57/26/17 37/51/12Mean (SD) LVEF, % 34 (10) 37 (11) 27 (7)Diabetes, % 14 11 18Treatment, %Frusemide 84 36 75Beta-blockers 71 64 79ACE inhibitor 78 79 77ARB 7 4 12Spironolactone 32 16 59a BMI, body mass index; LVEF, left ventricular ejection fraction; ACE, angio-tensinogen-converting enzyme; ARB, angiotensin receptor blocker.Clinical Chemistry 51, No. 7, 2005 1275counts and basic urine analysis. All were asymptomaticand underwent a complete examination by a cardiologist,including a visit, standard 12-lead electrocardiogram,Doppler echocardiographic examination, and a bicyclestress test (when older than 50 years), which excludedsilent heart disease. The median ACCESS and TRIAGEBNP concentrations were 10.5 ng/L (range, 1–53 ng/L;97.5th percentile, 45 ng/L) and 8.9 ng/L (range 5 to 52ng/L; 97.5th percentile, 35 ng/L), respectively. The distri-bution of BNP values measured by the 2 methods in thehealthy group is shown in Fig. 1 of the Data Supplementthat accompanies this Technical Brief at http://www.clinchem.org/content/vol51/issue7/.To evaluate the diagnostic accuracy of the 2 methods, wemeasured only samples from the 193 patients with a clini-cally ascertained diagnosis of heart failure. The clinicalcharacteristics of these patients are reported in Table 1.Cardiac morphology and function were assessed by 2-di-mensional echocardiography, or cardiac catheterizationwhen needed. The median (range) ACCESS and TRIAGEBNP concentrations measured were 171 (2–4833) ng/L and118 (5 to 4930) ng/L, respectively. The distributions ofBNP values measured by the 2 methods in these patientsgrouped according to the severity of heart failure [New YorkHeart Association (NYHA) functional classes I to IV] areshown in Figs. 1 and 2 of the online Data Supplement.However, the results obtained with both the TRIAGE andACCESS indicated a significant difference between the meanBNP values in healthy persons vs heart failure patients (P0.0001 by Scheffe` test after ANOVA).We used ROC curve analysis to evaluate the diagnosticaccuracy of the 2 methods in differentiating betweenhealthy persons and patients with heart failure. We foundno difference in diagnostic accuracy between the 2 meth-ods for differentiating healthy persons from patients withmild (NYHA class I and II; n  122; P  0.196) or severe(NYHA class III and IV; n  71; P  0.697) heart failure.For the TRIAGE, the areas under the curves were 0.840(SE, 0.027; 95% CI, 0.788–0.893) for patients with milddisease and 0.998 (SE, 0.002; 95% CI, 0.995–1.000) forpatients with severe heart failure, whereas for theACCESS system, the areas under the curves were 0.870(SE, 0.023; 95% CI, 0.825–0.916) for patients with milddisease and 0.997 (SE, 0.002; 95% CI, 0.993–1.000) forpatients with severe heart failure. The BNP values corre-sponding to a sensitivity of 95% in differentiating healthypersons from patients with mild heart failure were 7.5ng/L (corresponding specificity, 40%) for the ACCESSand 5.1 ng/L for the TRIAGE (corresponding specificity,29%). The BNP values corresponding to a specificity of95% were 41 ng/L (corresponding sensitivity, 65%) for theACCESS and 29 ng/L (corresponding sensitivity, 63%) forthe TRIAGE (Fig. 3 of the online Data Supplement).However, it is important to emphasize that the 95%sensitivity with the TRIAGE was obtained at a cutoff nearthe assay detection limit. The decision cutoff values fromthe current study are strictly related to our specific clinicalsetting, comparing healthy persons and patients withclinically ascertained heart failure. In routine clinicalpractice, several groups of individuals/patients suspectedof having a specific disease are usually compared.The present data confirm and extend previous resultssuggesting that BNP results are method dependent and thata single predefined common cutoff value cannot be used(5, 6). Furthermore, we demonstrated that immunoassaysthat use the same antibodies and calibration materials do notautomatically give the same results. However, the cutoff andreference values of these 2 methods are similar.References1. Clerico A, Emdin M. Diagnostic accuracy and prognostic relevance of themeasurement of the cardiac natriuretic peptides: a review [Review]. ClinChem 2004;50:33–50.2. Galvani M, Ferrini D, Ottani T. Natriuretic peptides for risk stratification ofpatients with acute coronary syndromes [Review]. Eur J Heart Fail 2004;6:327–33.3. Doust JA, Glasziou PP, Pietrzak E, Dobson AJ. A systematic review of thediagnostic accuracy of natriuretic peptides for heart failure. Arch Intern Med2004;164:1978–84.4. Clerico A, Del Ry S, Giannessi D. Measurement of cardiac natriuretichormones (atrial natriuretic peptide, brain natriuretic peptide, and relatedpeptides) in clinical practice: the need for a new generation of immunoassaymethods [Review]. Clin Chem 2000;46:1529–34.5. Clerico A, Prontera C, Emdin M, Passino C, Storti S, Poletti R, et al. Analyticalperformance and diagnostic accuracy of immunometric assays for themeasurement of plasma B-type natriuretic peptide (BNP) and N-terminalproBNP. Clin Chem 2005;51:445–7.6. Rawlins ML, Owen WE, Roberts WL. Performance characteristics of fourautomated natriuretic peptide assays. Am J Clin Pathol 2005;123:439–45.7. Apple FS, Panteghini M, Ravkilde J, Mair J, Wu AHB, Tate J, et al., on behalfof the Committee on Standardization of Markers of Cardiac Damage of theIFCC. Quality specifications for B-type natriuretic peptide assays. Clin Chem2005;51:486–93.8. Storti S, Prontera C, Emdin M, Passino C, Prati P, Fontani G, et al. Analyticalperformance and clinical results of a fully automated MEIA system for BNPassay: comparison with a POCT method. Clin Chem Lab Med 2004;42:1178–85.9. National Committee for Clinical Laboratory Standards. Evaluation of preci-sion performances of clinical chemistry devices: approved guideline. NCCLSDocument EP5-A, Vol. 19, No 2. Wayne, PA: NCCLS, February 1999:1–43.10. Bland JM, Altman DG. Applying the right statistics: analysis of measurementstudies. Ultrasound Obstet Gynecol 2003;22:85–93.DOI: 10.1373/clinchem.2005.048496Invasive Trophoblast Antigen (Hyperglycosylated Hu-man Chorionic Gonadotropin) as a First-Trimester Se-rum Marker for Down Syndrome, Martin J.N. Weinans,1*Ulrich Sancken,2 Raj Pandian,3 Jody M.W. van de Ouweland,4Henk W.A. de Bruijn,4 Jozien P. Holm,1 and Albert Mantingh1(1 Antenatal Diagnosis Unit, Department of Obstetricsand Gynaecology, and 4 Department of Pathology andLaboratory Medicine, University Hospital, Groningen,The Netherlands; 2 Institut fu¨r Humangenetik der Univer-sita¨t Go¨ttingen, Go¨ttingen, Germany; 3 Quest DiagnosticsNichols Institute, San Juan Capistrano, CA; * addresscorrespondence to this author at: Department of Obstet-rics and Gynaecology, University Hospital, PO Box 30.001,9700 RB Groningen, The Netherlands; fax 31503611806,e-mail martinweinans@planet.nl)Hyperglycosylated human chorionic gonadotropin (hCG)is a variant of hCG with more asparagine (N)-linked1276 Technical Briefs

Comparison of a fully automated immunoassay with a point-of-care testing method for B-type natriuretic peptide

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