Five days serum glucose stability at room-temperature in centrifuged fast-clotting serum tubes and the comparability with glucose in heparin-plasma and plasma containing citrate-stabilizer.

Glucose measurement plays a central role in the diagnosis of gestational diabetes mellitus (GDM). Because of earlier reports of overestimation of glucose in the widely used tubes containing granulated glycolysis inhibitor, the study assessed the performance of fast-clotting serum tubes as an alternative sample for the measurement of glucose. Glucose concentration in fast-clotting serum was compared to lithium-heparin plasma placed in an ice-water slurry after sample collection and glucose stability at room-temperature was studied. Blood samples from 30 volunteers were drawn in four different types of tubes (serum separator tubes, fast-clotting serum tubes, lithium-heparin tubes and sodium fluoride, EDTA and a citrate buffer (NaF-EDTA-citrate) tubes, all from Greiner Bio-One). Lithium-heparin tubes were placed in an ice-water slurry until centrifugation in accordance with international recommendations and centrifuged within 10 min. After centrifugation, glucose was measured in all tubes (timepoint T0) and after 24, 48, 72, 96 and 120 h of storage at 20-22 °C. NaF-EDTA-citrate plasma showed significant overestimation of glucose concentration by 4.7% compared to lithium-heparin plasma; fast-clotting serum showed glucose concentrations clinically equivalent to lithium-heparin plasma. In fast-clotting serum tubes, mean bias between glucose concentration after 24, 48, 72, 96 and 120 h and T0 was less than 2.4%. All individual differences compared to T0 were less than 6.5%. The results fulfill the acceptance criteria for sample stability based on biological variation. Fast-clotting serum tubes can be an alternative for the measurement of glucose in diagnosis and management of GDM and diabetes mellitus, especially when prolonged transportation is necessary.

Evaluation of the Sentinel-FOB gold faecal immunochemical test for the presence of haemoglobin using the automated Roche Cobas 8000 system.

OBJECTIVES: Faecal immunochemical tests (FIT) for the detection of haemoglobin are widely used in the diagnostic pathway of colorectal cancer (CRC) in patients presenting with lower abdominal symptoms. Several quantitative immunochemical tests are available, however only a few tests are available on high throughput automated analyzers. Here we report the analytical and clinical evaluation of the Sentinel-FOB Gold assay on the Roche Cobas 8000 automated system. DESIGN: and Methods: The assessment of the analytical performance of the assay on the Roche Cobas analyzer comprised determination of imprecision, accuracy, limit of detection, linearity, carryover and prozone effect. In the clinical evaluation part of the study 163 patients presenting for coloscopy with lower abdominal symptoms were included in the study. Diagnostic accuracy and optimal cutoff of the FIT-assay were determined by comparing faecal haemoglobin (f-Hb) concentrations with coloscopy findings as reference. RESULTS: We confirmed good analytical performance of the assay on the Roche Cobas system. Patients with CRC had significant higher f-Hb concentrations than patients with non-advanced adenomas and patients with normal findings by coloscopy. When applying a cutoff of 10 µg/g the sensitivity and specificity were 96.2% and 51.8%, respectively for CRC and 84.6% and 57.7% for CRC and advanced adenoma. CONCLUSION: The Sentinel-FOB Gold assay is appropriate for application on the automated Roche Cobas 8000 analyzer. The assay shows good analytical performance on the system. With regard to early detection of significant colorectal diseases a cutoff of 10 µg/g seems to be optimal.

Hb Aalesund (HBA2: c.400_406delAGCACCG), an Unstable α-Globin Variant Found in a Norwegian Patient Causing Moderate Hemolytic Anemia and Falsely High Hb A1c Using Ion Exchange High Performance Liquid Chromatography.

A new unstable hemoglobin (Hb) variant, named Hb Aalesund, was detected during Hb A1c measurement in a patient with a nearly compensated hemolytic anemia. Sequencing of the α-globin genes revealed a 7 bp deletion in exon 3 of the HBA2 gene (HBA2: c.400_406delAGCACCG) (NM_000517.4) causing a frameshift and a premature termination codon (PTC) two positions downstream. Apparently, the transcript bypassed nonsense-mediated decay (NMD), and a truncated protein was translated. The unstable Hb variant presumably underwent rapid denaturation, as heterozygosity of Hb Aalesund was associated with mild hemolytic anemia. In addition, the Hb variant interfered with Hb A1c measurement by cation exchange high performance liquid chromatography (HPLC), causing a falsely high Hb A1c result when using the Bio-Rad D10™ Hemoglobin Analyzer fast Hb A1c Program.

Reference Interval and Status for Serum Folate and Serum Vitamin B12 in a Norwegian Population.

BACKGROUND: Deficiencies of folate and vitamin B12 lead to an elevated serum concentration of homocysteine which has been associated with many diseases including cardiovascular disease. Laboratory algorithms often include initial testing of serum folate and vitamin B12. Reference intervals for these vitamins can vary significantly among populations for which dietary intakes may be different. The aim of this study was to establish reference intervals in a Norwegian population and to assess the folate and vitamin B12 status related to reference intervals. METHODS: Blood samples were taken from 144 healthy volunteers aged 18 - 65 years. A questionnaire provided data of medication, medical history, vitamin supplementation, alcohol consumption, and use of oral contraceptives and others. Serum folate and vitamin B12 concentrations were measured on the Abbott Architect i2000. Reference values were calculated using the bootstrap method. Results of serum folate, vitamin B12, and homocysteine from 1190 individuals from regional primary health care centers were evaluated related to reference values and the proportion of individuals with deficiency was estimated. RESULTS: Mean serum concentrations of folate and vitamin B12 were 11.9 nmol/L and 328 pmol/L, respectively. Men were found to have statistically significant higher vitamin B12 concentrations than women. 95%-reference intervals were calculated to 5.2 - 29.2 nmol/L for folate and 133 - 595 pmol/L for vitamin B12. 1.1% of the study population has serum vitamin B12-concentrations < 133 pmol/L and 3.4% has serum folate concentrations < 5.2 nmoI/L. CONCLUSIONS: The serum reference intervals for folate and vitamin B12 for a healthy, not vitamin-supplemented adult population were determined from 144 subjects. The application of these intervals will assist in the evaluation of folate and vitamin status.

Heterophilic antibody interference in commercial immunoassays; a screening study using paired native and pre-blocked sera.

BACKGROUND: Heterophilic antibodies are still an important source of interference in immunoassays. We have conducted a screening study for interference in a panel of commercially available assays using two sera known to contain high titer Fc-reactive heterophilic antibodies. METHODS: The sera were distributed to laboratories participating in the Nordic External Quality Assessment cooperation (EQANord). Duplicate samples pre-blocked with aggregated murine monoclonal MAK33 were also supplied. Discrepancies (>50%) between the results for native and blocked samples were used to classify the tested assays as susceptible to interference. A total of 170 different assay kits covering 91 analytes were tested. RESULTS: We found that 21 assays, covering 19 different analytes, were susceptible to interference from the heterophilic antibodies in the two sera. Many of these are clinically and commercially important assays. Some of the false results were grossly elevated and could have been detrimental to patient care in a clinical setting. CONCLUSIONS: Heterophilic antibodies with Fc-reactivity remain a threat. A more widespread use of antibody fragments and aggregated immunoglobulin could potentially improve the heterophilic antibody resistance of assays intended for clinical use.

Hepatic expression of A disintegrin and metalloproteinase (ADAM) and ADAMs with thrombospondin motives (ADAM-TS) enzymes in patients with chronic liver diseases.

BACKGROUND/AIMS: ADAMs (A Disintegrin And Metalloprotease) are multifunctional, membrane-bound and soluble cell surface glycoproteins with numerous functions in cell physiology. We assessed the expression of ADAMs in fibrotic liver disease of different aetiologies and clarified whether the expression of ADAMs is related to histological staging of fibrosis. In addition, the expression of ADAMs was determined in different cell types of liver. METHODS: Seventy-one biopsy samples from patients with chronic liver diseases were analyzed for mRNA expression of ADAM-8, -9, -12, -28, -TS1, -TS2, matrix metalloprotease (MMP)-2, -9 and tissue inhibitor of metalloproteinases-1 and -2 by quantitative real-time RT-PCR. RESULTS: The ADAM expression in liver injury is independent of aetiology. A strong correlation between ADAM -9, -28, -TS1 versus MMP-2 and SMA was identified. Activated hepatic stellate cells (HSC) showed increased mRNA expression of ADAM-8, -9, -12, -28, -TS2 compared to quiescent HSC. Significant differences between histological stages of fibrosis were found for ADAM-28, MMP-2 and MMP-9. CONCLUSIONS: The results suggest that ADAMs are differentially expressed in the liver. We assume that ADAM-9, -TS1 and -TS2 play a crucial role in extracellular matrix remodeling during fibrotic processes in the liver.

Drug screening in urine by cloned enzyme donor immunoassay (CEDIA) and kinetic interaction of microparticles in solution (KIMS): a comparative study.

Two commercially available drug-screening assays were evaluated: the Roche kinetic interaction of microparticles in solution (KIMS) assay and the Microgenics cloned enzyme donor immunoassay (CEDIA). Urine samples from known drug-abuse patients were analyzed for amphetamines, barbiturates, benzodiazepines, benzoylecgonine, cannabinoids, LSD, methadone and opiates. Samples with discordant findings for the two assays were analyzed by gas chromatography/mass spectrometry (GC/MS) or gas chromatography/electron capture detection (GC/ECD). Amphetamines showed 96.0% concordant results, with two false positive findings by CEDIA, three by KIMS and a further two false negatives by KIMS. Barbiturates showed 99.4% concordant results, with one false negative by KIMS. Benzodiazepines showed 97.4% concordant results, with two false negatives by KIMS (cutoff 100 microg/L, CEDIA cutoff 300 microg/L). Benzoylecgonine showed 17.8% concordant positive and 82.2% concordant negative results and no false finding by either assay. Cannabinoids showed 99.3% concordant results, with one sample negative by KIMS at a cutoff of 50 microg/L and positive by CEDIA (cutoff 25 microg/L). For LSD, 6.7% of findings were not in agreement. Methadone showed 97.5% concordant results, with two false positives by CEDIA, and one false positive and one false negative by KIMS. Opiates showed 96.9% concordant results, with no false KIMS results, but four false positives by CEDIA. The results indicate that the agreement of the CEDIA and KIMS results for the eight drugs is rather good (93.3-100%).

The membrane proteases adams and hepsin are differentially expressed in renal cell carcinoma. Are they potential tumor markers?

PURPOSE: ADAMs (a disintegrin and metalloproteinases) as cell surface proteins with adhesion and protease activity, and hepsin as a transmembrane protease have important roles in many biological processes. We assessed the expression of various ADAMs and of hepsin in human renal cell carcinoma (RCC), and correlated expression with clinicopathological data. MATERIALS AND METHODS: mRNA expression of ADAM-8, 17, 19, 28, TS1 and TS2, and of hepsin was investigated in paired tissue samples from cancerous and noncancerous parts of the kidneys of 27 patients with RCC who underwent tumor nephrectomy. Measurements were performed by quantitative real-time reverse transcriptase-polymerase chain reaction on a LightCycler instrument (Roche Applied Science, Mannheim, Germany). The data were related to housekeeping gene porphobilinogen deaminase mRNA. RESULTS: All ADAMs except ADAM-TS1 were significantly higher but hepsin was less expressed (at least p <0.05) in cancerous vs matched noncancerous tissue. Expression was differentially related to tumor stage. ADAM-8 and ADAM-TS2 over expression as well as decreased hepsin expression were associated with shorter patient survival. The Cox proportional hazards regression model revealed that ADAM-TS2 was an independent prognostic factor for cancer related death. ADAM-8 was the best predictor of distant metastases. CONCLUSIONS: The differential expression of hepsin and ADAMs suggests early and late involvement of membrane proteases in the development of RCC. Their association with the clinical outcome illustrates their potential usefulness as biomarkers for RCC.

Increased mRNA expression of ADAMs in renal cell carcinoma and their association with clinical outcome.

ADAMs (a disintegrin and metalloproteinase) are cell-surface proteins with adhesion and protease activity which play important roles in many biological processes. Little is known about their role in cancer. The aim of the study was to assess the quantitative expression of the ADAMs in human renal cell carcinoma (RCC) and to associate expression levels with clinicopathological data. We investigated the mRNA expression of ADAM-8, -17, -19, -28, ADAM-TS1, and ADAM-TS2 in paired tissue samples from cancerous and non-cancerous parts of the kidneys of 27 patients with RCC who underwent tumour nephrectomy. Measurements were performed by means of the quantitative real-time RT-PCR on a LightCycler instrument. ADAM-8, -17, and -19 were significantly higher expressed (p<0.05 at least) in cancerous compared with the matched non-cancerous tissue in pT1 and > or =pT2 tumours, ADAM-28 and ADAM-TS2 only in pT1 tumours, and ADAM-TS1 was not differently expressed. All ADAMs except ADAM-TS1 showed an increase of expression in the non-cancerous tissue with rising pT stage suggesting an early involvement of ADAMs in the development of RCC. The expression of ADAM-8 was related to a shorter survival of patients and was the best predictor of distant metastases. Our results indicate a potential role for ADAMs in RCC and that the overexpression might be a useful predictive tool.