Comparison of Five Common Analyzers in the Measurement of Chemistry Analytes in an Authentic Cohort of Body Fluid Specimens.

OBJECTIVES: Interpretation of body fluid (BF) results is based on published studies and clinical guidelines. The aim of this study is to determine whether the assays from five common commercial vendors produce similar results in BFs for 12 analytes in a BF cohort. METHODS: BFs (n = 25) and serum (n = 5) were analyzed on five instruments (Roche cobas c501, Ortho 5600, Beckman AU5800 and DXI800, Siemens Vista 1500, and Abbott Architect c8000) to measure albumin, amylase, total bilirubin, cholesterol, creatinine, glucose, lactate dehydrogenase (LDH), lipase, total protein, triglycerides, urea nitrogen, and carcinoembryonic antigen. Deming regression and Bland-Altman analysis were used for method comparison to Roche. RESULTS: Results were significantly different from Roche for LDH and lipase on Ortho and lipase on Siemens but similar for both BFs and serum. BF differences were larger than serum differences when measuring creatinine, glucose, and urea nitrogen on Ortho and glucose on Siemens. CONCLUSIONS: Five instruments used to perform BF testing produce results that are not significantly different except for lipase and LDH measurements. Bias of similar magnitude observed in both BF and serum should not affect interpretation. Further investigations into Ortho and Siemens measuring glucose and Ortho measuring creatinine and urea nitrogen are warranted.

Simultaneous determination of paracetamol and ciprofloxacin in biological fluid samples using a glassy carbon electrode modified with graphene oxide and nickel oxide nanoparticles.

A simple and highly selective electrochemical method using a glassy carbon electrode (GCE) modified with graphene oxide (GO) and nickel oxide nanoparticles (NiONPs) was developed for the simultaneous determination of paracetamol (PAR) and ciprofloxacin (CIP). The electrochemical characterisation of the modified GCE was performed by cyclic voltammetry and electrochemical impedance spectroscopy. The morphological characterisation of the GO and NiONPs was performed by scanning electron microscopy and transmission electron microscopy. Under optimised conditions, using square wave voltammetry, the simultaneous determination of PAR and CIP using the NiONPs-GO-CTS: EPH/GCE sensor shows a linear concentration range from 0.10 to 2.9µmolL-1 and 0.040-0.97µmolL-1, with detection limits of 6.7 and 6.0 nmol L-1, respectively. The NiONPs-GO-CTS: EPH/GCE sensor showed good reproducibility, repeatability and stability. Furthermore, the proposed method was successfully applied for the simultaneous determination of PAR and CIP in synthetic biological fluid samples.

Paper-based analytical device for instrumental-free detection of thiocyanate in saliva as a biomarker of tobacco smoke exposure.

This work describes a fast and simple assay for in situ detection of thiocyanate, i.e., a biomarker of tobacco smoke exposure, in human saliva. The assay is based on the formation of an iron(III)-thiocyanate colored complex in a paper-based sensing platform and subsequent image analysis using a scanner as detection device. Experimental parameters influencing the color intensity of the complex were fully evaluated, including the selection of detection conditions, type of paper substrate, test zone dimensions and composition as well as the stability of the paper-based device. Under optimal conditions, the detection limit was 0.06mM of thiocyanate, and the repeatability, expressed as relative standard deviation, was 3%. The proposed method, characterized by its simplicity, portability and low sample consumption, was applied to the detection of thiocyanate in a series of human saliva samples. Average thiocyanate levels in the ranges 0.28-0.87mM and 0.78-4.28mM were found for non-smokers and smokers, respectively. Recovery studies were carried out at two concentration levels, showing recovery values in the range of 96.1-103.6%.

Evaluation of the Lipid Interference for Siemens BN ProSpec Cystatin C Assay Using Pediatric Samples.

Endogenous interferents (lipids, hemoglobin and bilirubin) are a common cause of pre-analytical laboratory errors. We evaluated the effect of lipemia on Siemens cystatin C assay using pediatric samples. Lipemic samples were prepared by adding various concentrations of triglycerides into low and high cystatin C sample pools. Cystatin C concentrations were then measured on Siemens BN ProSpec analyzer and change of the analyte concentrations was determined. Low and high cystatin C sample pools were not affected by additions of lower lipid concentrations (150, 500 and 750 mg/dL), while the negative bias of <10% was seen with additions of higher lipid concentrations (1000 and 1500 mg/dL). Our results suggest that the BN ProSpec assay is a good alternative for cystatin C measurements in our pediatric population with no major interference from lipemia.

Clinical evaluation of the Nanoduct sweat test system in the diagnosis of cystic fibrosis after newborn screening.

UNLABELLED: After a positive newborn screening test for cystic fibrosis (CF), a sweat test is performed to confirm the diagnosis. The success rate of the generally acknowledged methods (Macroduct/Gibson and Cooke) in newborns varies between 73 and 99%. The Nanoduct sweat test system is easier to perform and less sweat is needed. The main aim of this study was to measure the success rate of the Nanoduct compared to current approved sweat test methods in a newborn population. After informed consent of the parents, newborns with a positive screening test for CF were included. The Macroduct or Gibson and Cooke and Nanoduct were performed in all infants, during the same appointment. The chloride concentration was determined by standard coulorimetry; conductivity was measured directly and converted to a NaCl molarity. One hundred eight newborns were included: 17 with CF, 7 with cystic fibrosis transmembrane regulator (CFTR)-related metabolic syndrome (CRMS), and 84 healthy children. The success rate of the Nanoduct was 93% and for the Macroduct/Gibson and Cooke 79% (McNemar, p = 0.002). The Nanoduct detected the same CF patients as the Macroduct/Gibson and Cooke; one CF patient had an equivocal result for both tests, and no patients were missed. The area under the receiver operating characteristic curve for detection of CF with the Nanoduct was 0.999, with ideal cutoff levels of 91 and 66 mmol/l, comparable to former studies. CONCLUSION: The success rate of the Nanoduct to collect sufficient sweat in infants was higher compared to the Macroduct and Gibson and Cooke.

Multidimensional gas chromatography methods for bioanalytical research.

Multidimensional gas chromatography (MDGC) methods are high-resolution volatile chemical separation techniques, and comprise classical heart-cutting MDGC and its more recent incarnation, comprehensive 2D GC. Although available for a long period, MDGC approaches are still not widely practiced in the field of bioanalysis, possibly reflecting the general preference for regular GC versus MDGC approaches. With the recent introduction of '-omic' techniques that emphasize global nontargeted profiling of metabolites within living systems, it is evident that MDGC is gaining momentum as a separation tool, since it offers very high resolution. By untangling metabolites within highly complex biological matrices, and expanding the metabolic coverage, MDGC plays a frontline role in '-omics' based studies. This review highlights state-of-the-art MDGC approaches, and summarizes the recent developments in bioanalytics.

Pediatric population reference value distributions for cancer biomarkers and covariate-stratified reference intervals in the CALIPER cohort.

BACKGROUND: Cancer biomarkers are commonly used in pediatrics to monitor cancer progression, recurrence, and prognosis, but pediatric reference value distributions have not been well established for these markers. The Canadian Laboratory Initiative on Pediatric Reference Intervals (CALIPER) sought to develop a pediatric database of covariate-stratified reference value distributions for 11 key circulating tumor markers, including those used in assessment of patients with childhood or adult cancers. METHODS: Healthy community children from birth to 18 years of age were recruited to participate in the CALIPER project with informed parental consent. We analyzed serum samples from 400-700 children (depending on the analyte in question) on the Abbott Architect ci4100 and established reference intervals for α-fetoprotein (AFP), antithyroglobulin (anti-Tg), human epididymis protein 4 (HE4), cancer antigen 125 (CA125), CA15-3, CA19-9, progastrin-releasing peptide (proGRP), carcinoembryonic antigen (CEA), squamous cell carcinoma antigen (SCC), and total and free prostate specific antigen (PSA) according to CLSI C28-A3 statistical guidelines. RESULTS: We observed significant fluctuations in biomarker concentrations by age and/or sex in 10 of 11 biomarkers investigated. Age partitioning was required for CA153, CA125, CA19-9, CEA, SCC, proGRP, total and free PSA, HE4, and AFP, whereas sex partitioning was also required for CA125, CA19-9, and total and free PSA. CONCLUSIONS: This CALIPER study established a database of childhood reference intervals for 11 tumor biomarkers and revealed dramatic fluctuations in tumor marker concentrations between boys and girls and throughout childhood. In addition, important differences between the adult and pediatric population were observed, further highlighting the need for pediatric-specific reference intervals.

Prognostic relevance of viable circulating tumor cells detected by EPISPOT in metastatic breast cancer patients.

BACKGROUND: Detection of circulating tumor cells (CTC) in breast cancer patients is currently performed in many clinical trials, using different technologies, in particular the EpCAM-dependent CellSearch® system. The purpose of this study was to investigate the incidence and prognostic relevance of viable CTC in a large cohort of metastatic breast cancer (MBC) patients. METHODS: A total of 254 MBC patients were enrolled in a prospective multicenter study at first diagnosis of metastatic disease or disease progression (before the start of a new treatment regimen). After EpCAM-independent enrichment, viable CTC releasing cytokeratin-19 as an epithelial cell marker were detected in the peripheral blood by an EPISPOT assay, and the Food and Drug Administration cleared CellSearch was used as the reference method. RESULTS: Using the EPISPOT assay, CTC were detected in 59% of MBC patients. The overall survival (OS) was linked with the CTC status measured by EPISPOT (P = 0.0191), which allowed stratification of MBC patients in low- and high-risk groups. This stratification could be improved by addition of the CTC status assessed by the CellSearch system. In multivariate Cox proportional-hazards regression analysis, the 3 methods used to determine the level of CTC (EPISPOT, CellSearch, and combination of EPISPOT/CellSearch) were compared by the Bayesian information criterion method. Interestingly, the combination of the EPISPOT and CellSearch assays was the strongest predictor of OS (hazard ratio, 22.6; 95% CI, 2.8-184.08). CONCLUSIONS: This is the first study in which CTC detection using the EPISPOT assay was evaluated on a large cohort of MBC patients, showing prognostic relevance of the presence of viable CTC.

Simultaneous determination of polyamines in human nail as 4-(N,N-dimethylaminosulfonyl)-7-fluoro-2,1,3-benzoxadiazole derivatives by nano-flow chip LC coupled with quadrupole time-of-flight tandem mass spectrometry.

BACKGROUND: Polyamines are active biogenic amines which play an important role in cell growth and proliferation and the synthesis of proteins and nucleosides. In recently studies, rapid tumor growth has been associated with markedly altered polyamine biosynthesis and accumulation. Therefore, the accurate detection and identification of all the polyamines simultaneously in a single analysis is becoming more and more important for the study of their biochemical roles. METHODS: The development of a simultaneous determination method for 9 polyamines in human nails was attempted by the combination of nano-flow chip LC and quadrupole time-of-flight tandem mass spectrometry (Q-TOF-MS/MS). The proposed method was for the determination in the nails of healthy persons and cancer patients. RESULTS: The derivatives of the polyamines in human nail were completely separated by a gradient of an 18 min duration wash program using a reversed-phase chip column. The polyamine derivatives were then introduced into the Q-TOF-MS/MS instrument and sensitively detected in the ESI(+) mode. Polyamine concentration was different based on the gender, i.e., PUT and SPD is higher in men than in women in the healthy persons. Additionally, in the lung cancer patients group, as compared with the healthy persons, concentrations of PUT, N¹-actPUT, and SPM were significantly increased. CONCLUSIONS: We here present a novel sensitive, simple method for the simultaneous determination of polyamines in human nails. Furthermore, we show that polyamines (ORN, DAP, CAD, N¹-actPUT, N8-actSPD, N¹-actSPM) were detected from the human nails. Human nails may may serve as the noninvasive bio sample for diagnosis of the chronic disease.

Development of des-gamma-carboxy prothrombin (DCP) measuring reagent using the LiBASys clinical analyzer.

BACKGROUND: Des-gamma-carboxy prothrombin (DCP), Lens culinaris agglutinin-reactive alpha-fetoprotein ratio (AFP-L3) and total alpha-fetoprotein (AFP) are tumor markers useful for diagnosing and determining the prognosis of hepatocellular carcinoma (HCC). There is a real need for measurement of these three markers on a one-assay platform. METHODS: A method of DCP measurement in human serum was developed using liquid binding assay (LBA), which enables rapid antigen-antibody reaction and bound/free separation on the LiBASys clinical analyzer. RESULTS: The dilution curve for DCP was linear up to 500 ng/mL. The limit of detection of DCP concentration was 0.5 ng/mL. Intra- and inter-assay coefficients of variation of DCP were 0.7%-2.4% and 2.2%-6.5%, respectively. This method was free from interference by hemoglobin, bilirubin, ditaurobilirubin, intrafat, ascorbate, galactose, glucose and rheumatoid factor. The analytical recoveries of DCP added to serum were 91.7%-108.2%. DCP concentration measured with the LBA method was linear and was significantly correlated with that measured with the ELISA method. CONCLUSIONS: The LiBASys clinical analyzer made possible measurement of the complementary tumor markers, HCC, total AFP, AFP-L3 and DCP.

Analytic evaluation of the beta-human chorionic gonadotropin assay on the Abbott IMx and Elecsys2010 for its use in doping control.

OBJECTIVES: The principal objective of this study was to compare the analytical performance of the Elecsys2010 (Roche Diagnostics) system with the IMx (Abbott laboratories) system for beta-hCG assay in order to assess its possible utility as a confirmation test for the quantitative measurement of beta-hCG in urine for doping control purposes. DESIGN AND METHODS: Urine samples with spiked standard known concentrations of beta-hCG and different urine samples from athletes were used in order to determine the calibration curve stability and linearity, detection limit, total, within-run and between-run precision, and method comparison for the IMx and Elecsys2010 systems for beta-hCG assay, along with the stability of samples, at room temperature and at 4 degrees C. RESULTS: The IMx assay was linear up to 500 IU/L, whereas the Elecsys2010 assay was linear up to 1000 IU/L. The detection limit for the IMx and Elecsys2010 systems were 0.75 IU/L and 0.25 IU/L, respectively. The total precision of the IMx and Elecsys2010 systems were

Automation and validation of a fast method for the assessment of in vivo oxidative stress levels.

BACKGROUND: The d-ROMs test for the evaluation of serum hydroperoxides (HP) is simple, reliable, and cheap. Furthermore, it can easily be adapted to automated analyzers. Changing from the manual to an automated procedure allows the simultaneous processing of a large number of samples in a greatly reduced time, avoiding manual handling of samples and reagents and reducing variability sources. METHODS: This study was performed to adjust the manual procedure to a routine automated method in the clinical laboratory. We carried out the d-ROMs test in sera from 90 subjects of both sexes (34 men and 56 women) with age ranging from 20 to 80 years (mean 51+/-14 years). All subjects were free from acute or chronic inflammatory disease, immunological disease and history or evidence of malignancy. Subjects were not on vitamin and/or antioxidant therapies. RESULTS: The detection limit of the assay was 40 AU. Linearity was observed up to 475 AU. The recovery ranged between 97% and 105%. Within- and between-run imprecision was <5%. The mean HP value was 304+/-8 AU, with no significant difference between men (291+/-10 AU) and women (311+/-11 AU). A significant positive correlation was observed between age and HP in the whole population (r=0.4, p=0.0002). CONCLUSIONS: The automated test for the estimation of serum hydroperoxides represents a reliable and feasible procedure for increasing efficiency and reducing costs compared to the manual method, and is particularly suitable for evaluating oxidative stress in a variety of clinical conditions.

Sample to sample carryover: a source of analytical laboratory error and its relevance to integrated clinical chemistry/immunoassay systems.

BACKGROUND: Integrated systems that combine clinical chemistry and immunoassay analyzers are used routinely. Sample to sample carryover is an inherent risk and can cause erroneously high patient test results for immunoassays. IVD manufacturers and laboratories must be aware of this phenomenon and guard against it. METHODS: We used a sample carryover protocol that directs the clinical chemistry module to process samples with very high immunoassay analyte concentrations followed by samples with very low concentrations for the same analyte. Low concentration samples were then tested by the immunoassay module to determine if the clinical chemistry module caused primary sample tube to primary sample tube carryover of the immunoassay analyte. RESULTS: Sample carryover was assessed on the Abbott ci8200 for HBsAg, AFP, beta-hCG, and PSA. Observed HBsAg carryover met the design specification of <0.1 ppm. Carryover for the other analytes was <0.1 ppm or below the assay limit of detection. CONCLUSIONS: IVD manufacturers must design integrated systems to minimize primary specimen tube carryover and avoid analytical laboratory error that can impact patient safety. Carryover testing is difficult for clinical laboratories to perform in order to verify system performance. Laboratories must consider the potential for specimen carryover and its impact on results whether moving primary sample tubes between separate analyzers or using an integrated system.

Urothelial cancer biomarkers for detection and surveillance.

Cancer is a complex process, and the US cancer-specific death rate has not changed in the last 50 years. Cure of the disease usually results from early diagnosis and treatment. Urothelial carcinoma (UC) has the highest recurrence rate of any cancer and is the second most common cancer of the genitourinary tract. It usually does not present at a metastatic stage, but only 50% of patients treated with cystectomy survive > or =5 years. There is a UC surveillance protocol, which includes cystoscopy, imaging, and cytology, to detect progression and allow early treatment of life-threatening UC. Many patients may not complete the surveillance protocol, and the cost of these studies is increasing. In addition, questions about the efficacy of these modalities have been raised. Therefore, bladder urinary tumor markers have been developed to aid in the diagnosis and surveillance of UC. Because urothelial cells are bathed in the urine and are continually shed, UC presents a unique opportunity to monitor bladder neoplasia in a noninvasive fashion. Currently, there are many research bladder tumor markers, but only a few are commercially available US Food and Drug Administration (FDA)-approved products. The commercially available bladder tumor markers and potential future markers are discussed. The ideal urinary bladder tumor test is still unavailable, but the eventual "gold standard" will consist of multiplex assays that analyze nucleic acids and proteins for detection. In addition, these tests would also reveal to the clinician both prognostic information and therapeutic targets for personalized medical treatment.

Clinical analysis by microchip capillary electrophoresis.

Clinical analysis often requires rapid, automated, and high-throughput analytical systems. Microchip capillary electrophoresis (CE) has the potential to achieve very rapid analysis (typically seconds), easy integration of multiple analytical steps, and parallel operation. Although it is currently still in an early stage of development, there are already many reports in the literature describing the applications of microchip CE in clinical analysis. At the same time, more fully automated and higher throughput commercial instruments for microchip CE are becoming available and are expected to further enhance the development of applications of microchip CE in routine clinical testing. To put into perspective its potential, we briefly compare microchip CE with conventional CE and review developments in this technique that may be useful in diagnosis of major diseases.

Differentiation between cerebrospinal fluid and serum with electronic nose.

OBJECTIVE: The study investigates the ability of the electronic nose to differentiate between cerebrospinal fluid (CSF) and serum and to identify an unknown specimen as CSF or serum. STUDY DESIGN AND SETTING: CSF and serum specimens were heated and tested with an organic semiconductor-based Cyranose 320 electronic nose (Cyrons Sciences, Pasadena, CA). Data from the 32-element sensor array were subjected to principal component analysis to depict differences in odorant patterns. RESULTS: The electronic nose was able to distinguish between CSF and serum isolates with Mahalanobis distance >5. Furthermore, the electronic nose was able to place unknown specimens in the appropriate class of CSF or serum. CONCLUSIONS: The electronic nose is a novel method that may allow rapid, low cost, and reliable distinction between CSF and serum in a clinical setting. SIGNIFICANCE: Because the results are available almost immediately, the electronic nose is a powerful tool that in the future may allow for rapid diagnosis of CSF leaks in the office setting.

Evaluation of analytical methods and workflow performance of the Architect ci8200 integrated serum/plasma analyzer system.

BACKGROUND: The Architect ci8200 is an integrated serum analyzer for photometric, electrochemical and immunological assays. Several assays of each category and the workflow performance of the system were compared with established laboratory procedures in two laboratories. METHODS: Measurements were compared with the ELECSYS 2010 (Roche Diagnostics) for CEA, PSA, FPSA, AFP, folate, vitamin B12, with the CENTAUR (Bayer) for TSH, T4, FT4, FSH and Estradiol, with the LIAISON (DiaSorin) for TSH, FT4 and FT3, with the Behring Nephelometer BN II (Dade-Behring) for ferritin, and with the INTEGRA 800 (Roche Diagnostics), and the AU640 (Olympus) for clinical chemistry assays. Workflow studies were performed to compare times of analysis required for defined analytical workloads. RESULTS: The coefficients of variation (CVs) for within-run imprecision were between 3% and 6% for CEA, PSA, FPSA, AFP and ferritin, and between 3% and 11% for TSH, FT4, FT3, folate and vitamin B12. The CVs for day-to-day imprecision for immunoassays were between 3% and 10%, except for vitamin B12 (CVs 11-13%) and FT4 (CV 10% -13%). For clinical chemistry tests corresponding CVs for within-run imprecision were < 1%, except for HDL, triglyceride, creatinine, ALT, LD and lipase (CVs<2%) and bicarbonate (CV 3%-6%) and magnesium (CV < 3%). The CVs for day-to-day imprecision for clinical chemistry tests were < 1%, except for sodium, CO(2), magnesium, phosphorus, glucose, uric acid, HDL, triglyceride, ALT, AST CK, lipase with CVs < 6% and for CO(2)<11%. Dilutional linearity testing of seven immunoassays and five clinical chemistry analytes resulted in recovery rates of 90-110%. Correlation studies with 15 immunoassays and 25 clinical chemistry tests showed acceptable agreements with established methods. Work flow analyses demonstrated a net gain in time of analysis up to 109 min depending on the size of the sample batch analyzed with the Architect ci8200 as the main analyzer as compared to the currently installed routine laboratory equipment. Median turn-around times were 7 and 30 min for chemistry assays and immunoassays, respectively, when ordered as STAT analyses, and 18 min when chemistry assays were ordered as routine determinations. CONCLUSIONS: Assays on the Architect ci8200 performed well, fulfilling quality control requirements as defined for instance by German quality control guidelines (RiliBAK). Method comparisons showed acceptable agreements with established assays. Workflow studies using the Architect ci8200 documented shorter times of analyses as compared with the conventionally established laboratory routine demonstrating the potential of integrated chemistry/immunoassay analyzers to provide faster and more efficient performance.

A importância das medidas de pressão arterial e da velocidade da onda de pulso no desenvolvimento da hipertrofia ventricular esquerda e no espessamento médio-intimal de carótidas em pacientes idosos / The importance of arterialpressure measurement and of pulse wave velocity n the development ao left ventricular hypertrophy and in carotid intima-media thickening in elderly patients

Objetivo: O objetivo deste estudo foi avaliar três grupos de indivíduos idosos, estratifcados segundo a MAPA em: normotenso (Grupo 1);provavelmente hipertenso sistólico isolado (Grupo II); e provavelmente hipertenso sisto-diastólico (Grupo III), na tentativa de identificar, entre as variáveis estudadas, aquelas que se correlacionavam com o determinismo das LOA definidas como HVE e ESPCED.Métodos: foram selecionados 89 indivíduos que se submeteram a exames laboratoriais, ao ECG com o objetivo de se excluir doença coronariana evidente, e à MAPA para a estratificação dos grupos. As variáveis analisadas para essas correlações compreenderam as medidas de PA casual e ambulatorial; as medidas de ESPCE e/ou ESPCD, através de ultr-som de carótidas; as medidas de MVE e IMVE, através da ecocardiografia; e medidas da VOP.Resultados: Os três grupos apresentaram distribuição semelhante em relação à idade, ao sexo, e aos índices antropométricos, bem como em relação à análise das médias dos parâmetros bioquímicos. Demostrou-se distribuição similar de ESPCED e de HVE, nos dois grupos de hipertensos, porém, com valores estatisticamente superiores em relação ao grupo de normotensos. As variáveis analisadas com significado estatístico e correlação positiva com a MVE foram: PAS24h, PP24h, PASVIG, PASSONO, PADSONO, e VOP; e a de correlação negativa foi a DESCPAS. As de correlação positiva com o ESPCED foram: PAS24h, PP24h, VS24h, PASVIG, PASSONO e VOP; e as de correlação negativa foram: DESCPAS e DESCPAD. Analisando o determinismo das LOA, observa-se que a PAS24h foi a única variável que manteve associação com a HVE(p igual0,0161), e a VOP com o ESPCED(p igual 0,033).Conclusões: demonstrou-se que a análise das variáveis de PA e VOP são de extrema validade na investigação de LOA em indivíduos idosos