Cautionary observations on preparing and interpreting brain images using molecular biology-based staining techniques

Though molecular biology-based visualization techniques such as antibody staining, in situ hybridization, and induction of reporter gene expression have become routine procedures for analyzing the structures of the brain, precautions to prevent misinterpretation have not always been taken when preparing and interpreting images. For example, sigmoidal development of the chemical processes in staining might exaggerate the specificity of a label. Or, adjustment of exposure for bright fluorescent signals might result in overlooking weak signals. Furthermore, documentation of a staining pattern is affected easily by recognized organized features in the image while other parts interpreted as "disorganized" may be ignored or discounted. Also, a higher intensity of a label per cell can often be confused with a higher percentage of labeled cells among a population. The quality, and hence interpretability, of the three-dimensional reconstruction with confocal microscopy can be affected by the attenuation of fluorescence during the scan, the refraction between the immersion and mounting media, and the choice of the reconstruction algorithm. Additionally, visualization of neurons with the induced expression of reporter genes can suffer because of the low specificity and low ubiquity of the expression drivers. The morphology and even the number of labeled cells can differ considerably depending on the reporters and antibodies used for detection. These aspects might affect the reliability of the experiments that involves induced expression of effector genes to perturb cellular functions. Examples of these potential pitfalls are discussed here using staining of Drosophila brain.     

Dot pattern generation technique using molecular dynamics

We have developed a new technique for generating homogeneously distributed irregular dot patterns useful for optical devices and digital halftoning technologies. To introduce irregularity, we use elaborately designed sequences called low-discrepancy sequences instead of pseudorandom numbers. We also use a molecular-dynamics redistribution method to improve the distribution of dots. Our method can produce arbitrary density distributions in accordance with a given design. The generated patterns are free from visible roughness as well as any moiré patterns when superimposed on other regular patterns. We demonstrate that our method effectively improves luminance uniformity and eliminates moiré patterns when used for a backlight unit of a liquid-crystal display.     

Uncertainties in estimated body burdens of caesium-137 by whole-body counting

It is very important to evaluate the uncertainties in individual monitoring for internal exposure of workers. The uncertainties in estimated body burdens of 137Cs with the JAERI whole-body counter were investigated using Monte Carlo simulation and measurements. It was found that the uncertainties of estimated body burdens with the whole-body counter are strongly dependent on various sources of uncertainty, such as radioactivity distribution within the body and counting statistics and that the 137Cs body burden assessed from the result of the whole-body count can be within +/- 60% in error.     

A novel ATP regeneration system using polyphosphate-AMP phosphotransferase and polyphosphate kinase

Polyphosphate-AMP phosphotransferase (PAP) and polyphosphate kinase (PPK) were used for designing a novel ATP regeneration system, named the PAP-PPK ATP regeneration system. PAP is an enzyme that catalyzes the phospho-conversion of AMP to ADP, and PPK catalyzes ATP formation from ADP. Both enzymes use inorganic polyphosphate [poly(P)] as a phosphate donor. In the PAP-PPK ATP regeneration system, ATP was continuously synthesized from AMP by the coupling reaction of PAP and PPK using poly(P). Poly(P) is a cheap material compared to acetyl phosphate, phosphoenol pyruvate and creatine phosphate, which are phosphate donors used for conventional ATP regeneration systems. To achieve efficient synthesis of ATP from AMP, an excessive amount of poly(P) should be added to the reaction solution because both PAP and PPK consume poly(P) as a phosphate donor. Using this ATP generation reaction, we constructed the PAP-PPK ATP regeneration system with acetyl-CoA synthase and succeeded in synthesizing acetyl-CoA from CoA, acetate and AMP. Since too much poly(P) may chelate MG2+ and inhibit enzyme activity, the Mg2+ concentration was optimized to 24 mM in the presence of 30 mM poly(P) in the reaction. In this reaction, ATP was regenerated 39.8 times from AMP, and 99.5% of CoA was converted to acetyl-CoA. In addition, since the PAP-PPK ATP regeneration system can regenerate GTP from GMP, it could also be used as a GTP regeneration system.     

A practical stress analysis for predicting fatigue limit of metal with axisymmetric complex surface

In order to predict the fatigue limit of a specimen with an axisymmetric complex surface, a practical method to estimate a stress concentration factor (SCF) of its surface was proposed. The roughness is coarse-grained by removing high frequency components and approximated with a parallel row of a local notch and innumerable average notches. Then, the notches are each approximated with the elliptical holes in the infinite plate, and the SCF is calculated approximately by superposing the elastic solutions of the holes. Moreover, FEM analyses were carried out on the various notch models which consist of the local notch and innumerable average notches to examine the application limit of the present method. Then, the validity of the application limit was examined by using the real roughness and the infinite parallel row of the various notches, and it was shown that the present method was available for the real roughness.     

Molecular simulation for adsorption of chlorinated hydrocarbon in zeolites

Equilibrium and isosteric heat of adsorption for the system of chloroform and USY-type zeolite were studied. The USY-type zeolite (PQ Co., SiO₂/Al₂O₃=70) was used both as a pure crystalline powder and as granulated particles with binder. Chloroform was reagent grade. The adsorption equilibria were measured using a gravimetric method and were expressed as isotherms. A chromatographic method (i.e. pulse response of chloroform through the USY column with helium carrier) was used to get the initial slope of the isotherms. In the simulation, the GCMC method was used to calculate amounts adsorbed for various conditions. FF parameters were confidently applied. And modified structure model was effective for simulation. This paper was presented at The 5th International Symposium on Separation Technology-Korea and Japan held at Seoul between August 19 and 21, 1999.     

Suppression of interface degradation in InGaAsP inP buried heterostructure lasers

In InGaAsP/InP buried heterostructure (BH) lasers, the degradation of BH interface between first- and second-growth step layers can be suppressed by employing the melt back process just before the second-step layer growth. It is confirmed this burying process give more reliable BH lasers than the conventional burying process. From the viewpoint of BH interface degradation, lasers lasing at 1.5 μm, where the melt back process naturally occurs during BH formation, are found to be more reliable than those lasing at 1.3 μm.