Diagnosis of active tuberculosis using MPB64, a specific antigen of Mycobacterium bovis.

Because the incidence of tuberculosis (TB) is still high in developing countries, an inexpensive and rapid diagnostic test for this infection is needed. To develop a screening test for TB, MPB64 antigen was produced by recombinant technology and purified with a polyhistidine tag. Next, serum and urine samples from patients with TB and uninfected individuals were examined by the dot-blot assay method using this purified antigen. Serum samples from patients with TB reacted more strongly with MPB64 antigen than did those from uninfected individuals. In addition, serum samples from TB patients with active infection reacted more strongly with the antigen than did samples from patients with inactive TB. When urine samples were assessed using this assay, similar results were obtained. Correlations between the data obtained from serum and urine samples were analyzed for all subjects, including uninfected individuals, and a strong positive correlation between the results of serum and urine tests (n = 36, r = 0.672) was found. The sensitivity and specificity of this assay for serum samples was 85.7 % and 85.0 %, and for urine samples 75.0 % and 85.0 %, respectively. These results suggest that dot-blot assay with MPB64 antigen could be a useful screening test for active TB. Because urine samples can be obtained more easily than serum samples and because urine is less contagious, urine testing should probably be employed for screening purposes.

A pseudohypoparathyroidism type Ia patient with normocalcemia.

Pseudohypoparathyroidism type Ia (PHP-Ia), one of 4 types of PHP, is a genetic disease characterized by clinical hypoparathyroidism caused by parathyroid hormone (PTH) resistance. In addition, patients with PHP-Ia show resistance to other hormones as well as Albright's hereditary osteodystrophy (AHO), a constellation of features including short stature, obesity, brachydactyly, ectopic ossifications, and/or mental retardation. Hypocalcemia is one of the hallmarks of PHP-Ia, but several PHP-Ia patients have been described to have normocalcemia. We encountered a 10-year-old girl with typical Albright's hereditary osteodystrophy with round face, short stature, brachydactyly, and obesity. Biochemical examination showed normocalcemia and increased PTH levels. Ellsworth-Howard test did not show any responses of urinary cAMP and phosphate. Based on these findings, she was diagnosed as having PHP-Ia with normocalcemia. Sequencing analysis of the GNAS gene identified a heterozygous missense mutation in exon 13 (R385H), which was previously reported in a PHP-Ia patient. The exact reason for her normocalcemia is not determined, but we must recognize heterogeneous biochemical findings even in PHP-Ia.

Development of quality evaluation sensor for fish freshness control based on KI value.

Recently, raw fish, sashimi, is becoming a popular dish in countries other than Japan. Therefore, in order to assure that the raw fish and shellfish are safe for human consumption, a quality evaluation sensor, which shows, at a glance, the quality of sashimi, was developed. The proposed sensor is based on the principle that the freshness of sashimi, which is judged from the KI value, can be determined from the degree of color change of thiazole blue (MTT: 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) due to the redox reaction of MTT accompanying the oxidation of hypoxanthine (Hx) by xanthine oxidase (XOD). The proposed sensor consists of 5 ml of 80% ethanol-1 M Tris-HCl buffer (pH 7.8) containing 2.0 mg of Hx, 2.0 mg of MTT and 0.38 unit of XOD. The proposed sensor and fish were kept together at 5, -10 and -20 degrees C, and the freshness of sashimi stored at each temperature was determined from the color change of the sensor. The concept "freshness of sashimi" can be expressed as remaining of validity (RDV), which is described in our previous study. A good relationship was obtained between the KI value and the RDV determined by the proposed sensor. From these results, the proposed sensor system can be used to non-destructively determine the fish freshness and RDV.

Over-expression of bHLH genes facilitate neural formation of mouse embryonic stem (ES) cells in vitro.

Mouse embryonic stem (ES) cells are useful tools for investigating differentiation into neurons and glial cells in vitro. In order to induce ES cells to differentiate into neural cells, many researchers have investigated the efficiency of induction. Embryoid body (EB) formation and retinoic acid are potent differentiation inducers known to be a trigger at the early stage of development. Basic helix-loop-helix (bHLH) is one of the important transcription factors, which is essential for premature neural formation. In NeuroD2 and Mash1-transfected cells, neural formation was observed at day 6 after the plating of embryoid bodies in culture. Nestin was detected in NeuroD2- and Mash1-transfected cells at day 10, and strong signal was detected in Mash1 transfectants by RT-PCR analysis. Map2 and Nurr1 were also detected strongly at the early stage in transfected cells compared with the wild type control, especially in the Mash1 transfectant. In immunocytochemical analysis, Tuj1-positive neurons were detected at high frequency in Mash1 transfectants and some cells were stained by tyrosine hydrogenase (TH), a marker of dopaminergic neurons. These results demonstrate that bHLH has a potential activity at an early stage for ES cells and can induce effective and rapid neural differentiation in vitro.