The automation of sediment urinalysis using a new urine flow cytometer (UF-100)

The UF-100 analyser is a fully automated instrument that stains the DNA and the membranes of the formed elements in native urine. The sample then passes as a laminar flow through a laser beam and light scattering, fluorescence and impedance are measured. The main purpose of the present work was to assess the analytical performance and the accuracy of the measurements of the UF-100 analyser. No carryover was observed, while the linearity was higher then the upper limit (40000 total particles microl(-1)) suggested by the manufacturer. The within-run imprecision was low, ranging from 17.7 % to 2.4% and was up to threefold better than manual microscopy. Comparison of results obtained by sediment microscopy (performed according to National Committee for Clinical Laboratory Standards (NCCLS) recommendations) and by the UF-100 analyser showed a linear correlation with r = 0.833 for erythrocytes, r = 0.934 for leukocytes, r = 0.880 for epithelial cells and r = 0.40 for casts. To evaluate the reliability of the UF-100 analyser in detecting bacteria we compared the results with the microbial culture (n = 608). Using a cut-off value of bacterial count above 1800 degrees l(-1) and at leukocyte count above 45 microl(-1), the analyser detected positive cultures with a sensitivity of 87 % and a specificity of 80 %. In conclusion, the UF-100 analyser can improve the work flow, increasing the output of urinalysis by reducing the number of specimens submitted for microscopy. Also the method provides reliable information for the identification of urinary tract inflammation and bacterial infection.

Interferon alfa-2a therapy in cryoglobulinemia associated with hepatitis C virus.

BACKGROUND: Essential mixed cryoglobulinemia is frequently associated with hepatitis C virus (HCV) infection. A beneficial effect of interferon alfa therapy has been reported, but we do not know whether the antiviral activity of the drug affects the clinical and biochemical manifestations of disease. METHODS: In a prospective randomized, controlled trial, we studied 53 patients with HCV-associated type II cryoglobulinemia. A group of 27 patients received recombinant interferon alfa-2a thrice weekly at a dose of 1.5 million units for a week and then 3 million units thrice weekly for the following 23 weeks. The 26 control patients did not receive anything apart from previously prescribed treatments. All patients were then followed for an additional 24 to 48 weeks. RESULTS: Interferon was usually well tolerated, but it was permanently discontinued in two patients because of atrial fibrillation and depression. Two of the 26 patients in the control group were lost to follow-up. After the treatment period, serum HCV RNA was undetectable in 15 of the remaining 25 patients who received interferon alfa-2a, but in none of the controls. In comparison with the control group, the 15 patients with undetectable levels of HCV RNA in serum had significant improvement in cutaneous vasculitis (P = 0.04) and significant decreases in serum levels of anti-HCV-antibody activity (P = 0.007), cryoglobulins (P = 0.002), IgM (P = 0.002), rheumatoid factor (P = 0.001), and creatinine (P = 0.006). After treatment with interferon alfa-2a was discontinued, viremia and cryoglobulinemia recurred in all 15 HCV RNA-negative patients. On resumption of treatment, three of four patients had a virologic, clinical, and biochemical response. CONCLUSIONS: The therapeutic efficacy of interferon alfa-2a in HCV-associated cryoglobulinemia is closely related to its antiviral activity, thus supporting the idea that HCV infection may be a cause of this disease.

Hepatitis C virus infection in patients with essential mixed cryoglobulinemia.

OBJECTIVE: To study the association between hepatitis C virus (HCV) infection and essential mixed cryoglobulinemia. SETTING: Wards and clinics of the Ospedali Riuniti di Bergamo and Ospedale di Treviglio e Caravaggio, Italy. PATIENTS: Fifty-one patients with essential mixed cryoglobulinemia associated with glomerulonephritis and 45 controls with noncryoglobulinemic glomerulopathies. MEASUREMENTS: Antibodies to hepatitis C virus (anti-HCV) in sera from patients with essential mixed cryoglobulinemia and from controls, using two enzyme-linked immunosorbent assays (c100 ELISA and c22/c200 ELISA) and a recombinant immunoblot assay (4-RIBA); cryoprecipitate anti-HCV before and after use of dithiothreitol, a substance able to destroy IgM antibodies with rheumatoid factor activity, in patients with essential mixed cryoglobulinemia; serum HCV RNA by polymerase chain reaction in patients with essential mixed cryoglobulinemia. RESULTS: In patients with essential mixed cryoglobulinemia, the c22/c200 ELISA detected anti-HCV in 98% of serum samples (95% CI, 90% to 100%), whereas the rate of reactivity remained at 2% (CI, 0% to 12%) in the control group (P less than 0.0001). These results were confirmed by the 4-RIBA in 66% of patients with essential mixed cryoglobulinemia. The study of cryoprecipitate by c100 ELISA showed anti-HCV in 41% (Cl, 28% to 56%) of patients. After dithiothreitol, the rate of reactivity increased to 94% (CI, 84% to 99%; P less than 0.0001 by the McNemar paired chi-square test), suggesting that the elimination of rheumatoid factor leads to unmasking of anti-HCV in cryoprecipitate. Polymerase chain reaction detected HCV RNA in 13 of 16 sera from patients with essential mixed cryoglobulinemia. CONCLUSIONS: The extremely high prevalence of anti-HCV in serum and cryoprecipitate along with the frequently associated serum HCV RNA suggests a close relation between essential mixed cryoglobulinemia and chronic HCV infection.

Measurement of alkaline phosphatase activity in serum with N-methyl-D-glucamine as a buffer: evaluation of the method for routine use.

Some aspects of the measurement of alkaline phosphatase activity concentration in human serum, using N-methyl-D-glucamine as a buffer, were evaluated with a view to the possible routine use of the method. The evaluated characteristics included: the temperature-dependence of the pH of the buffer; the effect of adding the magnesium/zinc ions buffer; the effect of the serum volume fraction; the substrate-starter versus the serum-starter mode; the effect of modifying the formulation of reagents; the within-run and the between laboratory imprecision; the correlation with an alternative routine method. Also, sex- and age-related reference values were produced, based on 2968 values from selected reference sample groups in 7 laboratories. In general, the results demonstrate the robustness of the method, its adaptability to a variety of mechanized analysers, and hence its feasibility as a routine measurement method.

Development and improvement of a commercial uric acid enzymatic determination kit on a centrifugal fast analyzer.

The performance of a uric acid determination kit has been evaluated for five months, under routine conditions, in a General Hospital Biochemical Laboratory. An anomalous increment in fresh serum blanks was noted in the kit when first introduced. This interference, probably due to alcohol dehydrogenase contamination, was corrected by addition of 50 mmol/l (NH4)2SO4 to the reagent. Results obtained with this modified reagent correlate perfectly with those obtained with modified kits by Smith Kline Instruments (SKI), and with many other determination methods. Correlations are discussed and explanations for differences in statistical data are offered. Within run and between run precision data are presented. The kit fits perfectly with the needs of centrifugal fast analyzers and discrete micro analyzers, on account of its speed, reliability and precision.