Volume 48, Issue 4

Therapeutic drug monitoring (TDM) is a multidisciplinary activity. Because laboratory reports are part of the patient's chart, some clinical information is required. In order to guarantee quality and safety, an increasing number of TDM departments have implemented a quality management system. The aim of the present article is to review the three phases of TDM: the pre-analytical, analytical and post-analytical phases. In the pre-analytical phase, it is necessary to acquire a valid specimen collected at the specific time window. Analytical methods should be validated, assessing possible interfering substances. The objective of the post-analytical phase is the final report, which should include correct interpretation, as well as possible advice. Appropriate pharmacokinetic interpretation avoids unnecessary costs and leads to clinical benefits. Clin Chem Lab Med 2010;48:437–46.

Background: Genotyping is crucial for the identification of genetic markers underlying the development of neoplastic diseases and for determining individual variations in response to specific drugs. Technologies which can accurately identify genetic polymorphisms will dramatically affect routine diagnostic processes and future therapeutic developments in personalized medicine. However, such methods need to fulfill the principles of analytical validation to determine their suitability to assess nucleotide polymorphisms in target genes. Approach: This article reviews recent developments in homogeneous technologies for the genotyping of single nucleotide polymorphisms. Here, homogeneous methods essentially refer to “single-tube” assays performed in a liquid phase. For the appropriate choice of any method, several criteria must be considered: 1) detection of known genetic variations; 2) analytical performance including specificity, sensitivity and robustness of the method; 3) availability of large platforms and required equipment; 4) suitability of platforms and tests for routine diagnostics; 5) suitability for high throughput implementation. Content: This review is intended to provide the reader with an understanding of these various technologies for pharmacogenomic testing in the routine clinical laboratory. A brief overview is provided on the available technologies for the detection of known mutations, a specific description of the homogeneous platforms currently employed in genotyping analysis, and considerations regarding the proper assessment of the analytical performance of these methods. Based on the criteria proposed here, potential users may evaluate advantages and limitations of the various analytical platforms and identify the most appropriate platform according to their specific setting and diagnostic needs. Clin Chem Lab Med 2010;48:447–59.

Background: Critical value reporting is considered an essential tool to ensure the quality of medical laboratory services. Important issues include defining cut-off values, assessing responsibility for communication and adopting information technology solutions to improve notification. Here, we report the state of critical value reporting in a large cohort of Italian laboratories and comparison with Q-Probes surveys from the College of American Pathologists as representatives of the US situation. Methods: To compare critical value policies and procedures, formulation of critical values list with critical values limits and monitoring tools, a web-based questionnaire was formulated for 389 institutions participating in the External Quality Assessment Schemes of Veneto Region, in Italy. Results: A total of 90 clinical laboratories submitted data. Accredited laboratories represented 82.2% of participants, but written procedures for reporting were indicated by 70.5% of participants. Relevant differences between US and Italian policies have been observed, particularly regarding who provides the notification and on the formulation of the cut-off threshold for critical values. Conclusions: Accreditation according to international standards can decrease differences regarding the management of critical values across laboratories of different countries. However, the issues concerning critical limits should be debated and a consensus critical values list should be considered. Automated systems could offer improvements regarding some issues, such as who makes the notification, reducing the time spent in notification of critical values. Surveys for comparing and improving existing policies regarding critical values should be promoted at an international level. Clin Chem Lab Med 2010;48:461–8.

Background: The Abbott RealTi m e hepatitis C virus (HCV) Genotype II (Abbott Molecular Inc.) for HCV genotyping, which uses real-time PCR technology, has recently been developed. Methods: Accuracy and sensitivity of detection were assessed using the HCV RNA PHW202 performance panel (SeraCare Life Sciences). Consistency with restriction fragment mass polymorphism (RFMP) data, cross-reactivity with other viruses, and the ability to detect minor strains in mixtures of genotypes 1 and 2 were evaluated using clinical samples. Results: All performance panel viruses were correctly genotyped at levels of >500 IU/mL. Results were 100% concordant with RFMP genotypic data (66/66). However, 5% (3/66) of the samples examined displayed probable genotypic cross reactivity. No cross reactivity with other viruses was evident. Minor strains in the mixtures were not effectively distinguished, even at quantities higher than the detection limit. Conclusions: The Abbott RealTi m e HCV Genotype II assay was very accurate and yielded results consistent with RFMP data. Although the assay has the advantages of automation and short turnaround time, we suggest that further improvements are necessary before it is used routinely in clinical practice. Efforts are needed to decrease cross reactivity among genotypes and to improve the ability to detect minor genotypes in mixed infections. Clin Chem Lab Med 2010;48:469–74.

Background: The identification of human mitochondrial DNA (mtDNA) sequence variations, especially single nucleotide polymorphisms (SNPs), is important for many applications. The PCR-ligase detection reaction (LDR) method can reduce false-positives and eliminate the need for both post-PCR and post-ligation purifications in SNP analyses. In addition, it has been successfully employed to detect point mutations in various nuclear genes. In this study, we used the PCR-LDR platform to characterize mtDNA SNPs. Methods: Multiplex PCR-LDRs were used to genotype 19 mtDNA single nucleotide polymorphic sites from 812 samples. Performance of the method was assessed by direct sequencing of 44 samples. Results: We established an overall 97.4% success rate with 99.2% accuracy using the multiplex PCR-LDR methodology. Conclusions: The PCR-LDR mtDNA genotyping technique is simple, highly accurate, has high-throughput, and is cost-effective. Therefore, this method is applicable to mtDNA haplotyping in various applications. Clin Chem Lab Med 2010;48:475–83.

Background: Blepharophimosis, ptosis, epicanthus inversus syndrome (BPES) is an autosomal dominant developmental disorder that includes an eyelid malformation associated with (type I) or without (type II) premature ovarian failure (POF). Mutations in the forkhead transcription factor 2 ( FOXL2 ) gene, a member of winged/forkhead transcription factor family, are responsible for both types of BPES. The purpose of this study was to identify mutations in FOXL2 in Taiwanese patients with BPES. Methods: The karyotype and genomic DNA was prepared from the leukocytes of peripheral venous blood samples. The coding and flanking region sequences of FOXL2 were analyzed by directed or cloning sequencing. Results: The karyotypes of these patients did not show significant variation, especially on the 3q23 region. Two mutations in FOXL2 were identified in two familial cases. One was c.855-871dup (17-bp insertion) associated with POF. The other was c.384G>A (TGG>TGA), a novel mutation that resulted in non-sense changes of the encoded protein, i.e., p.W128X. Conclusions: Our results expand the spectrum of FOXL2 mutations and confirm the mutation hotspot in FOXL2 in Taiwanese BPES patients. Clin Chem Lab Med 2010;48:485–8.

Background: Human β-defensins (hBDs) are small cationic, widely expressed proteins involved in innate immunity that exert strong bactericidal activity toward various pathogens. However, the role of hBDs in various diseases to which bacterial infection add severity, as it is in celiac disease (CD), is not yet clear. We analyzed the expression of the hBD1 , hBD2 , hBD3 and hBD4 genes in patients with CD during the active phase and after remission following a gluten-free diet to determine their role in development and relapse of CD. Methods: We studied 21 unrelated adults with CD (confirmed by anti-thyroglobulin antibodies and intestinal biopsy); 14 were evaluated at diagnosis, before diet modification, and seven after 2 years of a gluten-free diet. Thirty-six unrelated adults served as controls. We analyzed the mRNA expression of hBD1, 2, 3 and 4 on biopsy samples of duodenum obtained from all patients during endoscopy for diagnostic purposes. We used real-time polymerase chain reaction with TaqMan probes and obtained gene expression data using the delta-Ct method. Results: hBD1 mRNA was significantly lower in patients with active CD compared with patients on diet modification, whereas the mRNA levels of the other three defensins did not differ significantly between the two subgroups. Interestingly, the gluten-free diet restored only partially hBD1 expression as compared to a normal group of celiac-free subjects. Conclusions: Our data reinforce the evidence that hBD1 expression is greatly reduced in the duodenum of patients with active CD. It also strengthens the concept that reduced activity of immune peptides may predispose individuals to bacterial proliferation that contributes to the pathogenesis of CD. Clin Chem Lab Med 2010;48:489–92.

Background: Various biomarkers have been proposed to help diagnose inflammatory and infectious diseases. The goal of this study was to evaluate the effectiveness of some of these markers in children seen in an emergency service, and to compare a number of routinely used tests with measurements of serum amyloid A (SAA) and C-reactive protein (CRP) levels. Methods: A total of 83 children with clinically suspected inflammation (n=17) or infection (n=66) who had been seen at the emergency department were evaluated. Complete blood count, erythrocyte sedimentation rate (ESR), CRP (using two methods) and SAA measurements were performed. Results: There was no difference in the parameters studied between the inflammation and infectious groups. However, when the infectious group was subdivided into bacterial (n=21) and viral infections (n=43), significantly higher leukocyte, CRP and SAA values were observed in the former group. CRP was the most accurate indicator of inflammatory and infectious processes [Kappa concordance index (KCI)=0.89], followed by SAA (KCI=0.73) and ESR (KCI=0.73). The worst results were obtained for the immature/total neutrophil ratio (KCI=0.02). Conclusions: Measurement of CRP showed the best performance for indicating infectious or inflammatory processes. ESR and SAA had good concordance and yielded very similar results. CRP may be included in protocols for pediatric assessment in emergency departments. Clin Chem Lab Med 2010;48:493–9.

Background: Accurate identification of bacterial infections in patients presenting at the emergency department is crucial for early and rational antibiotic treatment. In this situation, using a cut-off of 0.25 μg/L for procalcitonin allows for carefully monitoring of febrile patients. Most previous studies have been performed with the reference B·R·A·H·M·S PCT KRYPTOR ® assay. The goal of this study was to compare this test with the VIDAS B·R·A·H·M·S PCT ® assay and to validate clinically relevant cut-off thresholds. Methods: This prospective study was conducted in adults presenting to the emergency departments of a tertiary hospital. We included 305 consecutive patients that had procalcitonin requested. Procalcitonin was measured first with the KRYPTOR, then with the VIDAS systems. Statistical analysis consisted in Passing and Bablok and Bland-Altman plots. Results: In the overall cohort, 176 patients had procalcitonin concentrations measured using both methods and were well correlated. The Bland-Altman plot exhibited a bias of 0.108 [95% confidence interval: –0.044 to 0.260]. The concordance at different procalcitonin cut-off thresholds, respectively of 0.1, 0.25, 0.5 and 2 μg/L, indicated that above 0.25 μg/L, the κ coefficient was >0.80. Conclusions: A highly significant correlation was observed between the two automated assays. Procalcitonin concentrations obtained from both methods led to the same clinical interpretation. Clin Chem Lab Med 2010;48:501–4.

Background: Recent advanced techniques in glycobiology have produced a number of tumor marker candidates. As a result from the glycomic approach, we found that fucosylated haptoglobin in sera was a possible tumor marker for pancreatic cancer (PC). Although Aleuria aurantia lectin (AAL) blotting can detect fucosylated haptoglobin, it is difficult to quantify fucosylated haptoglobin precisely. To overcome this problem, we developed a fucosylated haptoglobin detection kit as a sandwich enzyme-linked immune sorbent assay (ELISA) using AAL and the Fab portion of anti-haptoglobin antibody. In the present study, we investigated the clinical application of this lectin-antibody ELISA kit to measure fucosylated haptoglobin in PC. Methods: We measured fucosylated haptoglobin in patients with PC with a lectin-antibody ELISA kit. The fucosylated haptoglobin measured with this assay was compared with lectin blotting data, and the discrepancy was analyzed by immunoprecipitation methods. The concentration of fucosylated haptoglobin was investigated with respect to the clinical stage of PC. We also measured fucosylated haptoglobin, using 397 cases of several types of cancers including PC, benign diseases, and normal controls. Results: The sensitivity and specificity for the differential diagnosis of PC from normal controls was 50% and 91%, respectively. The results from lectin-antibody ELISA were significantly correlated with data from previous AAL blotting studies. Positive rates of fucosylated haptoglobin with this method in patients with PC were significantly higher in cases of stage IV compared with other clinical stages. Fucosylated haptoglobin was increased in several types of cancers, in which fucosylated haptoglobin was reported to increase. Conclusions: While certain cases showed a discrepancy in fucosylated haptoglobin concentrations between the lectin-antibody ELISA and conventional lectin blotting, this novel type of lectin-antibody ELISA might be useful for a tumor marker for PC. Clin Chem Lab Med 2010;48:505–12.

Background: To provide a more comprehensive clinic marker of tryptophan (TRP) catabolism in patients with systemic lupus erythematosus (SLE), we developed a simple and efficient method that simultaneously measured serum TRP, kynurenine (KYN), and kynurenic acid (KYNA) using high performance liquid chromatography with fluorescence detection (HPLC-FD). Methods: A simple and specific high performance liquid chromatography (HPLC) method was developed for simultaneously quantitative determination of TRP, KYN and KYNA with fluorescence detection (FD) using programmed wavelength and on-column fluorescence derivatization. Thirty patients with SLE and 80 healthy control subjects were analyzed for serum TRP metabolites using the assay we developed. The tryptophan breakdown index (TBI) and neuroprotective ratio (NPR) were calculated. Results: The retention time of KYN, KYNA and TRP were 8.5 min, 13.7 min and 17.6 min, respectively. The linear range for TRP was 0.245 ~ 196 μmol/L, the limit of detection (LOD) was 0.001 μmol/L and average recovery was 103.71%. The linear range for KYN was 0.049 ~ 98 μmol/L, the LOD was 0.0245 μmol/L, and average recovery was 97.45%. The linear range for KYNA was 1.05 ~ 2093 nmol/L, the LOD was 0.05 nmol/L, and average recovery was 100.60%. Inter-day and intra-day relative standard deviations (SDs) were <5%. Phenylalanine, tyrosine, 5-hydroxytryptamine and creatinine did not interfere with the method. The results showed great differences in TRP, KYN and KYNA contents and TBI between patients with SLE and healthy controls, but little difference in NPR. Conclusions: The method is simple, fast, accurate, and meets the requirements for simultaneous determination of TRP, KYN and KYNA in serum. Clin Chem Lab Med 2010;48:513–7.

Background: Random-access analyzers that employ dedicated probes may be affected by carryover. Sample carryover, involving analyte from one sample that is measured in a subsequent sample, is most often detected in measurement procedures that have wide reportable ranges. However, reagent carryover can be more difficult to detect as it may involve only one pair of measurement procedures. Methods: Our laboratory noted several patient samples with total cholesterol <2.58 mmol/L (100 mg/dL) during the initial months after a new chemistry analyzer was installed. The problem seemed to occur intermittently, approximately once per week. Results: Immediate reanalysis of affected samples resulted in measured values that were 1.03–2.58 mmol/L (40–100 mg/dL) higher. Evaluation of reagent carryover revealed a significant decrease in total cholesterol after analysis of creatine kinase (EC 2.7.3.2). Carryover disappeared when an additional reagent probe wash was applied, whereas the root of the problem was eliminated with replacement of the reagent probes. The insidious nature of reagent carryover made it difficult to initially detect the problem, which affected two of the three instruments in our laboratory. Conclusions: Laboratorians should be aware of the potential for carryover in random access analyzers, and should be mindful of appropriate troubleshooting techniques. Clin Chem Lab Med 2010;48:519–21.

Background: Prediction of the duration of motor block after injection of a local anesthetic into cerebrospinal fluid (CSF) would be a very useful tool in clinical practice. However, previous attempts have not shown conclusive results. In this work, glycorrhachia is demonstrated to be an adequate predictive parameter after spinal anesthesia using 0.5% hyperbaric bupivacaine. Methods: Two mL of local anesthetic through a continuous spinal catheter was administered to 40 patients. CSF was sampled at different time intervals from the onset of infusion to motor recovery. CSF bupivacaine concentrations were measured using chromatography. An automated analyzer was used for determining glycorrhachia in the same samples. Results: For all patients, good correlation (r 2 >0.95, p<0.05) was obtained. From these results, it was possible to develop a general model which establishes the relationship between CSF glucose and bupivacaine concentrations (R 2 =0.987, p<0.05). Motor block is reached when CSF glucose concentration is about 245 mg/dL (13.5 mmol/L), which corresponds to 35 μg/mL of bupivacaine. Conclusions: Glycorrhachia measured during surgical intervention in patients undergoing spinal anesthesia with hyperbaric bupivacaine provides a mechanism for predicting the duration of motor block in a rapid and simple manner. Clin Chem Lab Med 2010;48:523–30.

Background: Urine, being an ultrafiltrate of plasma, is a rich source for biomarker discovery. Since potential new disease markers are often present in low concentrations, a prefractionation/enrichment step could be useful in the discovery process. To enhance the detection of low-abundance proteins, three immuno-affinity depletion approaches were evaluated. Methods: To remove the most abundant proteins from a human urine sample, GenWay™ Spin IgY-12 kit, HPLC Agilent Hu-PL7 and a home-made column vs. human serum albumin [immuno-affinity column (IAC)] were compared. Quantification of total proteins, 2-D gel electrophoresis (2-DE), Progenesis gel images analysis and mass spectrometric proteins identification were applied to evaluate these strategies. Results: Reproducibility of depletion columns, by estimating protein content of unbound fractions, were: 343±20.0 μg, 5.8%; 186.3±13.3 μg, 7.2%; 292±20.6 μg, 8.8% [mean± standard deviation (SD), CV%], for GenWay™, Agilent and IAC methods, respectively. To isolate urinary protein after depletion, ethanol precipitation provided the highest recovery (80%). Applying 2-DE and Progenesis analysis, the number of spots visualized on the gels was 468±21, 331±7, 368±22 and 304±7 (mean±SD) for GenWay™, Agilent, IAC, and the undepleted urine pool sample, respectively, with a significant difference p<0.001 compared to the GenWay procedure. Conclusions: The sequential procedure of urine samples using multi-protein immuno-affinity depletion represents a valid tool for simplifying 2-DE analysis of the urine proteome. Particularly, the GenWay™ kit followed by ethanol precipitation was found to be the most efficient method for exploring the urine proteome. Clin Chem Lab Med 2010;48:531–5.

Background: Pregnancy-associated plasma protein-A (PAPP-A) has been associated with peripheral artery disease (PAD). The aim of this study was to evaluate the utility of PAPP-A as a marker for long-term mortality in patients with atherosclerotic PAD. Methods: PAPP-A serum concentrations were measured using an enzymatically amplified two-step sandwich-type immunoassay in 487 consecutive patients admitted to a tertiary care hospital with symptomatic PAD. The main outcome measure was all-cause mortality at 5 years. Results: During follow-up, 114 patients died and 373 survived. The median PAPP-A concentration was higher among decedents compared with survivors (0.96 vs. 0.78 mU/L, p=0.024). The area under the receiver operating characteristic curve for the prediction of 5-year mortality by PAPP-A was 0.57 [95% confidence interval (CI), 0.53–0.61; p=0.026]. Survival probability was not significantly associated with PAPP-A concentrations using Kaplan-Meier curve analysis. However, univariate Cox proportional-hazards regression analysis revealed that PAPP-A was associated with 5-year mortality [risk ratio 1.25; 95% CI, 1.05–1.50; p=0.013 per one standard deviation (SD) increase in log transformed values]. In the multivariate model using a bootstrapping method, the predictive value of PAPP-A remained significant (risk ratio 1.31; 95% CI, 1.01–1.73; p=0.024 per 1 SD increase in log transformed values), even after adjustment for clinical confounders and other biomarkers, such as high-sensitivity C-reactive protein and amino terminal pro-B-type natriuretic peptide. Conclusions: In this study, PAPP-A was an independent predictor of 5-year all-cause mortality in patients with symptomatic PAD. However, based on the weak association between PAPP-A and outcome in our cohort, we consider PAPP-A measurements to not be useful in clinical practice for prognostic purposes in patients with PAD. Clin Chem Lab Med 2010;48:537–42.

Background: Circulating concentrations of matrix metalloproteinase 9 (MMP-9) are associated with cardiovascular disease mortality in patients with coronary artery disease. We investigated the determinants of MMP-9 concentrations by analyzing MMP-9 genotypes and risk factors for cardiovascular disease. Methods: A total of 596 individuals were recruited for this study. Six single nucleotide polymorphisms (SNP) with coverage of the MMP-9 gene region were analyzed; and two genotypes, rs3918242-TT and rs2274756-AA, which were in nearly complete linkage disequilibrium, were associated with higher MMP-9 values (p=0.007 and p=0.008, respectively). In age- and gender-adjusted regression models, MMP-9 concentrations were positively associated with the homeostasis model assessment of insulin resistance (HOMA-IR) index and triglyceride concentrations, fasting serum insulin, fasting plasma glucose, C-reactive protein, fibrinogen, and serum amyloid A. By multivariate analysis, rs2274756 genotypes, age, smoking status, fibrinogen and fasting plasma glucose concentrations were all independently associated with MMP-9 (p=0.007, p=0.033, p=0.003, p=0.013, and p=0.012, respectively). Conclusions: The rs2274756-AA and rs3918242-TT genotypes, younger age, current smoking status and increased fasting plasma glucose, and fibrinogen concentrations were independently associated with high serum MMP-9 concentrations in Taiwanese individuals. Clin Chem Lab Med 2010;48:543–9.

Background: C-reactive protein (CRP) has been shown to contribute to the risk of cardiovascular disease. Several single nucleotide polymorphisms (SNPs) at the CRP locus have been found to be associated with CRP concentrations. We hypothesized that genetic variants associated with high CRP may confer a greater risk of stroke and recurrence. The hypothesis was tested in a case-control study and during follow-up of the case population. Methods: Two genetic variants associated with serum CRP in the CRP gene were genotyped in a Chinese case-control study comprised of 1572 stroke patients and 1485 controls. The case population was followed for a median of 4.5 years (range, 0.1–6.0 years). Results: During a median 4.5-year follow-up for 1526 stroke patients, 299 recurrent strokes were identified. No association was found between the SNPs or haplotypes of the CRP gene and stroke and its recurrence. Conclusions: Although these variants and corresponding haplotypes in the CRP gene are associated with serum CRP concentrations, our study does not support that variants and corresponding haplotypes studied here have a major influence on risk of stroke and stroke recurrence. Therefore, we speculate that CRP is not a causal factor for stroke. Clin Chem Lab Med 2010;48:551–4.

Background: Hyperhomocysteinemia is a risk factor for ischemic heart disease. Several other mechanisms apply also to dilative types of heart failure of various, non-ischemic etiologies. We hypothesized that hyperhomocysteinemia is associated with left ventricular (LV) dilatation and hypertrophy in dilative cardiomyopathy. Methods: Homocysteine was measured in 66 individuals with suspected cardiomyopathy. Cardiac magnetic resonance imaging was used to assess LV volume, mass, and wall stress. Results: Hyperhomocysteinemia (>12 μmol/L) was found in 45 patients (68%). LV mass was greater in these patients compared with individuals with normal homocysteine (83±27 vs. 67±19 g/m 2 ; p<0.02). Homocysteine was increased in patients with increased brain natriuretic peptide ≥100 pg/mL (18.3±5.9 vs. 14.9±5.1 μmol/L; p=0.018). LV mass, LV end-diastolic and end-systolic volume (LVEDV, LVESV) were significantly increased in individuals in the upper quartile compared with the lower quartile (90±25 vs. 65±18 g/m 2 , p=0.021; 114±50 vs. 71±23 mL/m 2 , p=0.042; 76±51 vs. 36±22 mL/m 2 , p=0.045). LV dilatation (LVEDV≥90 mL/m 2 ) was more common in hyperhomocysteinemia (>12 μmol/L, p=0.0166). Normalized LV mass was correlated with homocysteine (r=0.346, p=0.065). Homocysteine was not significantly correlated with LVEDV (r=0.229, p=0.065), LV end-diastolic wall stress (r=0.226, p=0.069) and LV ejection fraction. Conclusions: Hyperhomocysteinemia appears to be, at least in part, involved in a disproportional LV dilatation, where the ensuing hypertrophy is not sufficient to compensate for the increased wall stress. A potential mechanism is the hyperhomocysteinemia associated increase in oxidative stress that favors muscle fiber slippage. Clin Chem Lab Med 2010;48:555–60.

Background: Adiponectin, present in different multimeric forms in circulation, is increasingly used in clinical settings as a cardiometabolic marker. The development of several commercially available enzyme-linked immunosorbent assays (ELISA) has allowed for the widespread measurement of adiponectin in research as well as in clinical practice. The comparative performance of these assays is thus an issue of major relevance. Methods: The analytical performance of four different ELISAs (LINCO, B-Bridge, SPIbio and ALPCO) was evaluated. Samples from 102 cardiac patients and 40 healthy subjects were tested using LINCO and ALPCO. The latter is able to measure the concentrations of multimers. For the multimer assay, an error propagation study was performed. A subset of subjects was tested by all four ELISAs for comparison purposes. Results: Bland-Altman plots revealed differences between the results of the four ELISAs, mainly for ALPCO. However, a strong correlation between the different ELISAs (ALPCO, r=0.83; B-Bridge, r=0.98; SPIbio, r=0.91 vs. LINCO) was observed. Total adiponectin measured by LINCO showed a better association with cardiac disease [receiver-operating characteristic (ROC) curves] than total and high molecular weight adiponectin measured by ALPCO. Conclusions: The results of this study contribute to a better understanding of the methodological features of the several ELISAs, and help in the evaluation and comparison of the relative results. Clin Chem Lab Med 2010;48:561–8.