Polymorphisms of the genes for enzymes participating in the metabolism of several carcinogens have been identified. These polymorphisms are associated with varieties in the enzymatic activity or substrate specificity, and thus additive or multiplicative risks for development of various human cancers. In this study, we compared the genotype frequencies of four enzymes (CYP1A1, CYP2D6, N-acetyltransferase and glutathione S-transferase) from cervical cancer patients and healthy control individuals, to estimate the importance of polymorphisms in human papillomavirus infection and development of cervical cancer. The frequency of the Slow acetylator N-acetyltransferase genotype differed significantly between cervical cancer patients and healthy controls (P < 0.05, x² = 4.24). No significant difference was found between these two groups in the frequency of any of the other genotypes studied. These findings suggest that the arylamine compounds metabolized by N-acetylation play an important role in human cervical carcinogenesis.

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We established an enzyme-linked immunosorbent assay for tissue-unspecific alkaline phosphatase using specific monoclonal antibodies prepared against liver alkaline phosphatase. The concentration of serum tissue-unspecific alkaline phosphatase was determined in the range of 1.5 to 100 ng/ml. The coefficients of variation in the between-day assay and the within-day assay were 3.2-6.9% and 6.8-7.9%, respectively. The recovery of tissue-unspecific alkaline phosphatase from various serum samples was 94.1%-95.8%. This method can be clinically applied. In serum of a patient with infantile hypophosphatasia which is one of six types of hypophosphatasia that is deficient in tissue-unspecific alkaline phosphatase, both the concentration and the activity of serum tissue-unspecific alkaline phosphatase were lower than those in the sera of healthy individuals. This suggests that the lower activity of tissue-unspecific alkaline phosphatase in this patient with infantile hypophosphatasia is due to the lower expression of tissue-unspecific alkaline phosphatase, the production of the non-immunoreactive abnormal protein of tissue-unspecific alkaline phosphatase and/or less of it being transported from the cell membrane to the circulation.

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The myeloperoxidase activity of synovial fluid and elastase in plasma were examined in patients with rheumatoid arthritis (RA) or osteoarthritis (OA) and control subjects. The elastase activity was significantly higher in RA patients than control subjects (mean ± SD 345.1 ± 187.6pg/1 versus 143.1 ± 67.5μg/l ; p<0.001). The myeloperoxidase activity insynovial fluid was determined by using a guaiacol method with a fixed time rate assay. The myeloperoxidase activity were significantly higher in RA patients than OA patients (mean ±SD 224±134μ/l versus 18±15μ/l;p<0.02). These results indicated that myeloperoxidase is secreted from neutrophils together with elastase. In view of known formation of reactive oxidants by synovial neutrophils, we propose that an extracellular myeloperoxidase-H₂O₂ system plays an important role in tissue damage.

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The use of liquid type reagent is required in common use at clinical laboratories to keep more precise and accurate quality control. However, there are several problems concerning to the raw materials of reagents which contains enzymes, coenzymes and substrates etc. Because these are unstable in liquid solution for longterm storage. For the measurement of uric acid, we developed new thermostable recombinant uricase from Bacillus sp., TB9O strain. Although uricase from Candida sp. has been currently used for the determination of uric acid, the enzyme activity in a liquid solution is unstable for long-term storage. A recombinant enzyme from Bacillus sp., TB9O strain has two excellent properties, which make it superior to that from Candida sp.;the enzyme activity in a liquid solution has been kept for more than one year under the storage condition of 10°C, the enzyme has a broad optimum pH range. It is a suitable liquid reagent for the determination of uric acid. We evaluated a liquid type reagent using recombinant type enzyme for the determination of uric acid. A good linearity, precision and no influences of interfering substances were obtained. This liquid reagent with using a recombinant uricase is useful for the routine test at clinical laboratories.

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[in Japanese]

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We developed a method for measuring the total and alternative pathway activities of the complement system using rabbit γ-globulin (RGG)-coupled liposomes and anti-RGG antibody. The lysis of these liposomes in the present system as well as the hemolysis in the conventional method was expressed by von Krogh's equation. The reproducibility of the present method was satisfactory. The total activity and the activity of the alternative pathway measured by the present method were correlated to the activity measured by the conventional Mayer's method (r=0.555 and 0.351, respectively). Measurement of the complement activities in healthy subjects revealed that the total activities measured by the present method and the activities measured by Mayer's method increased with age, but the activities of the alternative pathway were independent of age. Furthermore, the total activities measured by the present method in diabetic subjects were lower than those in healthy subjects. However, there was no significant difference in the activities measured by Mayer's method between diabetic and healthy subjects. These findings, as well as the homogeneity of the liposomes and the simplicity of these preparation, suggested that the present method using liposomes could be a new clinical parameter for measurement of the complement activities.

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Binding capacities of immunoassay reagents for remnant-like particles, mixed gel suspension of anti-apo B-100 and apo A-I monoclonal antibodies coupled to Sepharose 4B, to LDL and HDL were examined by HPLC method. The resolution and sensitivity for detection of cholesterol (C) in serum lipoproteins have been improved using a new special column (TSK gel Lipopropak, 7.5 mm I. D., 600 mm length / TOSOH Co. Tokyo) as well as a new eluent (TSKeluent LP-1). Within-run precision of this method by applying 100 μl of highly diluted serum (× 1,001) was excellent both for LDL-C and HDL-C (CV value: 2.49% and 2.48 %). About 98% of LDL and HDL of serum specimens containing less than 210 mg/dl of LDL-C, and 100 mg/ dl of HDL-C was bound to the immunoassay reagent gel. Therefore, in the routine assay technique, in which 5 μl of serum was added to 300μl of mixed gel suspension and the reaction mixture was gently shaken for 60 min at room temperature, maximun binding capacities of the reagent gel for LDL-C and HDL-C were suggested to be 210mg/dl and 100mg/dl, respectively.

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We developed a high-performance liquid chromatographic method with ultraviolet detection using a column-switching system for the determination of vanilmandelic acid (VMA levels in human urine. The lower detection limit of VMA in urine was 0.5μg/ml. VMA in sample solutions was stable for at least 25 h at room temperature under a room light.
The recovery of VMA in urine was 92% over the range of 0.50 to 5.03μg/0.5ml urine. Reproducibilities (CV) for within- and betweenassays of samples containing three concentrations of VMA (low, medium and high) were 1.3%, 1.2% and 1.1% (n=10 each), and 5.0%, 3.1%, and 1.9% (n= 5 each), respectively. The VMA levels in urine measured by the present method (y) correlated well with those by a conventional method (x) r = 0.92, y 0.72x+0.19, n= 41).

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