Development of a plasma maltose assay method as a screening test for upper gastrointestinal disorders.

BACKGROUND AND OBJECTIVE: The disaccharide loading test is a method to assess gastric mucosal damage. Since Trelan-G75, which is used for the sugar tolerance test, contains disaccharide maltose, if maltose is detected at a high sensitivity in the sample blood used in the sugar tolerance test, screening for upper gastrointestinal mucosal damage can be made simultaneously with the sugar tolerance test for the diagnosis of diabetes. METHODS: Glucose-6-phosphate is generated by treating maltose with maltose phosphorylase, ß-phosphoglucomutase, and glucose-1,6-bisphosphate. Then, change in the absorbance at 405 nm is measured by the enzymatic cycling method using Thio-NADP, ß-NADPH, and Glucose-6-phosphate dehydrogenase. After evaluating the optimal condition for this method, it is mounted on an automatic biochemical analyzer, and samples after the sugar tolerance test were assayed. RESULTS: Regarding the performance of this method, the repeatability was 10-50 µmol/L with a CV of ≤1.1%. Concerning the assay range, a curve passing the origin with a range of linearity up to 120 µmol/L was obtained. No effect of dyes or sugars in the blood was noted. As a result of application to patients with gastric mucosal disorders (those who had a health checkup), significant differences were observed depending on the stage of atrophic gastritis. DISCUSSION: This method has a high sensitivity and a high precision and can be used for high-speed analysis on an automatic analyzer. It has the potential to be used as a screening test for gastric mucosal damage.

A survey of the reactivity of in vitro diagnostic bilirubin reagents developed in Japan using artificially prepared bilirubin materials: A comparison of synthetic delta, unconjugated, and taurine-conjugated bilirubin.

BACKGROUND: In vitro diagnostic bilirubin reagents based on oxidation with bilirubin oxidase or vanadic acid for total and direct-reacting bilirubin are widely used in Japan; however, their reactivity to unconjugated and conjugated bilirubin and delta bilirubin has not been completely disclosed by manufacturers. We used artificially prepared bilirubin materials to investigate the reactivity with four in vitro diagnostic bilirubin reagents. METHODS: Porcine unconjugated bilirubin solution, chemically synthesized ditaurobilirubin solution, and chemically synthesized delta bilirubin solution were used as surrogates of naturally occurring unconjugated bilirubin, conjugated bilirubin, and delta bilirubin, respectively. The total bilirubin and direct-reacting bilirubin concentrations were measured by three bilirubin oxidase methods and one vanadic acid method, and the observed concentrations were compared with those obtained by the diazo-based reference measurement procedure. RESULTS: The unconjugated bilirubin and delta bilirubin concentrations were similar when any of the four in vitro diagnostic bilirubin reagents were used during total bilirubin measurement. This was consistent with reference measurement procedure and exhibited a converged inter-method variation. Compared with reference measurement procedure, significantly low ditaurobilirubin concentrations were observed by the in vitro diagnostic bilirubin reagents despite the converged inter-method variation. In delta bilirubin measurement, some reagents reacted doubtfully with unconjugated bilirubin, while showed lower ditaurobilirubin concentrations than its corresponding total bilirubin concentration. Reactivity with delta bilirubin was different for each method including reference measurement procedure. Some reagents were developed to react less with delta bilirubin and others to strongly react with delta bilirubin. CONCLUSIONS: We revealed the reactivity of IVD-TB and IVD-DB reagents to artificially prepared bilirubin materials, and their consistency with reference measurement procedure. The delta bilirubin data results vary depending on the reagents used.

Lipoprotein(a)-cholesterol: a significant component of serum cholesterol.

BACKGROUND: Lipoprotein(a) [Lp(a)] is known to be a cholesterol-rich lipoprotein, however, the contribution of Lp(a)-cholesterol [Lp(a)-C] to the serum cholesterol and LDL-C levels has not yet been fully evaluated. METHODS: We determined the serum Lp(a)-C in 55 subjects with serum Lp(a) concentrations ranging from 9 to 129 mg/dl. To measure the serum Lp(a)-C concentrations, we developed an immunoaffinity gel assay; serum was incubated with Sepharose 4B gel coupled with immunoglobulin G (IgG) prepared from a polyclonal anti-Lp(a) goat antiserum. After separating Lp(a) from other lipoproteins, we determined the serum Lp(a)-C concentrations. Validation of the assay showed satisfactory results in terms of the specificity and reproducibility. RESULTS: The mean cholesterol content of Lp(a), determined as Lp(a)-C/Lp(a), was 29.5±10.4%. The serum Lp(a)-C values were found to be highly correlated with the serum Lp(a) mass (r=0.923, p<0.001). At serum Lp(a) levels of over 50mg/dl, the contribution of Lp(a)-C to the serum total cholesterol was 10.2%. Further, the Friedewald formula overestimated the serum LDL-C by 20.4%. CONCLUSIONS: Lp(a) contains approximately 30% cholesterol in each molecule. In subjects with markedly elevated serum Lp(a) concentrations, the Lp(a)-C values should be taken into account when evaluating the serum LDL-C.