Use of a test diet in screening for neuroblastoma reduces false-positives attributable to food intake.

We attempted to reduce false-positives during screening for neuroblastoma using a qualitative urine test by introducing a test diet without foodstuffs known to cause false-positive results. In preliminary in-vivo experiments, intake of contra-indicated foods such as orange juice or banana was shown to result in high percentages of false-positive results several hours after food intake. False-positive results were obtained even after 24 hours among breast-fed infants whose mothers received orange juice. In a controlled field trial the false-positive rate was reduced to 2.84% among 540 infants taking the test diet compared with 5.05% among 9,844 control infants following conventional guidance on contra-indicated foods (p less than 0.05). For comparison, a questionnaire survey of nationwide screening in Japan in 1987 revealed that 66% of the screening centres employed qualitative urine tests, either a Spot or Dip method. False-positive rates, including those due to inappropriate urine collection, ranged from 0.4% to 33.7% (mean 7.1%). Rates ranged from 0.2% to 18.7% (mean 3.4%) in the remaining 34% of screening centres employing a quantitative method with high performance liquid chromatography.

Preservation and transportation of bacteria by a simple gelatin disk method.

A wide range of bacterial species, e.g., Enterobacteriaceae and Neisseria, Streptococcus, Branhamella, Haemophilus, Gemella, Pseudomonas, Flavobacterium, and Bacteroides species, were successfully preserved for 1 to 5 years by our gelatin disk drying method. The beta-lactamase activity of penicillinase-producing Neisseria gonorrhoeae was retained for more than 3 years with this method. Good results were also obtained upon airmailing many strains of N. gonorrhoeae embedded in gelatin disks from Japan to the United States. Neisseria, Branhamella, Gemella, and Haemophilus organisms suspended in the reagent used in the preparation of the gelatin disks could be preserved for 6 to 12 months after freezing the cell suspensions at -20 degrees C. Furthermore, modification of our gelatin disk preservation method made possible the safe long- and short-distance transportation of clinical isolates. Our methods can be used by any small laboratory, since they require only conventional instruments and reagents.

Simplified purification and biophysicochemical characteristics of Kanagawa phenomenon-associated hemolysin of Vibrio parahaemolyticus.

Kanagawa phenomenon-associated hemolysin (K-hemolysin) was purified by Sephadex gel and ion-exchange column chromatography, after the culture supernatant had been adsorbed on and eluted from diethylaminoethyl-Sepharose CL-6B, and acid precipitated. K-hemolysin was a heat-stable and trypsin-susceptible protein with an apparent molecular weight of 44,000, the subunit of which was 22,000. The isoelectric point was 4.9. The minimum hemolytic dose was 0.1 mug/ml. The fifty percent lethal dose by intravenous injection was 1.4 mug. Electron microscopy of the small intestine of suckling mice orally challenged with the highest dose (50 mug) not only showed disappearance of epithelial cell microvilli, but also structural disturbances of the endoplasmic reticulum and mitochondrial swelling. One blueing dose representing permeability factor activity was 0.3 mug, and positive reaction in the rabbit ileal loop appeared at above 125 mug. Besides these data in experimental models, we discovered the appearance of an antibody in patients which neutralizes K-hemolysin during the course of the disease. This finding reinforces our view that K-hemolysin plays a most significant role in the pathogenesis of this enteric human disease.

Preservation of Neisseria gonorrhoeae by the gelatin-disc method.

Studies of Neisseria gonorrhoeae are difficult to perform because of the organism's poor survival in vitro. To solve this problem we tried to preserve the organism by a gelatin-disc method. The rate of survival and changes of variations in some biochemical properties of eight strains of N. gonorrhoeae were followed for three years. These studies proved that preservation was satisfactory with only a 1/10 reduction of the living cells. Another trial showed that the organism survived for over six months after being frozen at -20 degrees C. The colonial types, agglutination against red cells from rabbit and guinea pig, and antibiotic susceptibility to penicillin, chloramphenicol, tetracycline, kanamycin, and streptomycin did not change after three years' preservation.