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Biomedical subjects

Y Hamaguchi

Publications and source records attributed to Y Hamaguchi.

At least 145 records · Page 8Linked to original sources
PubMed1984

Antifibrinolytic activity in middle ear effusion.

No activity of the plasmin and plasminogen activators could be detected in middle ear effusion (MEE). A streptokinase proactivator, which was not plasminogen and markedly enhanced the activation of plasminogen by streptokinase, could be detected. MEE had an apparent inhibitory activity against both thiol protease (papain) and serine proteases (plasmin and trypsin). All these findings indicate that MEE contains unsaturated protease inhibitors and is in a state of predominance of inhibitors over fibrinolytic activity.

Antifibrinolytic Agents↗
PubMed1984

Duplicated immunoglobulin gamma 2a genes in wild mice.

The Japanese wild mouse, Mus musculus molossinus, has duplicated immunoglobulin gamma 2a genes, whereas most of the laboratory-strain mice have single gamma 2a genes. This duplication provides a unique system in which molecular mechanisms of the evolutionary rearrangement of the immunoglobulin gene can be studied. For this purpose it is important to elucidate the organization of the immunoglobulin genes surrounding the duplicated gamma 2a genes. It is also interesting to assess the distribution of this haplotype among the wild mouse population. Partially overlapping chromosomal segments that encompass the gamma 2b, gamma 2a-1, gamma 2a-2, and epsilon genes were isolated from phage libraries containing DNA fragments from M. m, molossinus. The organization of this region has been elucidated as 5'-gamma 2b-(18 kb)-gamma 2a-1-(19 kb)-gamma 2a-2-(14 kb)-epsilon-3'. Both gamma 2a-1 and gamma 2a-2 genes have their own switch regions in the 5' flanking region. Two homology units of 13 kb, each containing the gamma 2a gene, were identified. The homology units alternate with repetitive sequences of low homology, making the location of the recombination site obscure. Seventeen of 31 Japanese and Chinese wild mice screened had duplicated gamma 2a genes, indicating a wide distribution of this genetic event in the Asian wild mouse population. Moreover, three alleles each of the gamma 2b and gamma 2a loci have been observed in this small sample population. An unequal crossing-over event between heterologous haplotypes is proposed to account for the duplication of the gamma 2a gene, and a candidate for one member of the pair was identified among wild mouse haplotypes. These results suggest that the immunoglobulin heavy-chain gene family of the Japanese and Chinese wild mouse subspecies has accumulated not only polymorphic divergence due to point mutations, but also dynamic rearrangements such as gene duplication, during its recent evolution since divergence of these subspecies from the European subspecies about one million years ago.

Animals↗
PubMed1982

Effects of phalloidin microinjection and localization of fluorescein-labeled phalloidin in living sand dollar eggs.

Effects of microinjection of phalloidin on fertilization and cleavage of sand dollar (Clypeaster japonicus and Scaphechinus mirabilis) eggs were studied. The drug, previously injected into unfertilized eggs, showed no effect on the elevation of the fertilization membrane upon insemination up to an intracellular concentration of 50 microM. However, the movement of the egg pronucleus to the sperm pronucleus was inhibited and the fusion of pronuclei did not occur. The subsequent development no longer took place. When phalloidin was injected into fertilized eggs, the thickness of the cortical layer increased and the microvilli became conspicuous. Both nuclear division and cleavage were inhibited at the intracellular concentration of more than 20 microM, though the latter seemed to be more sensitive to phalloidin than the former. Fluorescein-labeled phalloidin (FL-phalloidin) was injected into eggs in order to investigate F-actin localization by fluorescence microscopy. In both unfertilized and fertilized eggs, FL-phalloidin was localized in the cortical layer within 1 min after injection. It was also localized in the cortical layer as radially oriented rod-like structures when injected into fertilized eggs before the disappearance of the nuclear membrane. No distinct fluorescence was detected in the mitotic apparatus or in the cleavage furrow. FL-phalloidin redistributed gradually into egg cytoplasm. In unfertilized eggs, fluorescent rods were found especially in the egg pronucleus 30 min after injection.

Actins↗
PubMed1982

Quantitation of nasal secretory IgA by enzyme-linked immunosorbent assay.

A simple and sensitive technique, enzyme-linked immunosorbent assay (ELISA) was used for the quantitation of secretory IgA (sIgA) in order to study the synthesis and transport of sIgA in the nasal secretion from patients with chronic sinusitis. Rough-surface glass beads coated with antisecretory component antibody were employed as the solid phase. The minimal detectable concentration of sIgA was 50 ng/ml in this study. sIgA averaged 226.3 +/- 217.4 mg/dl and total IgA averaged 302.0 +/- 137.5 mg/dl.

Albumins↗
PubMed1982

The lethal protein from Kintoki beans (Phaseolus vulgaris) identified as a lectin.

A glycoprotein isolated from Kintoki beans (Phaseolus vulgaris cultivar Kintoki) agglutinated human erythrocytes of all types and erythrocytes of rat, rabbit, sheep, and mouse. The lectin activity was not affected by 1 hr heating at 60 degrees C, but decreased slightly on heating for the same period at 70-80 degrees C and markedly at 90-100 degrees C. The activity was inhibited by galactose, lactose, N-acetyl galactosamine and fetuin. The inhibition was, however, weak, as often found for nonspecific lectins. The activity did not change when tyrosine residues or small parts of amino groups were modified, but decreased considerably when histidine residues or carboxyl groups were modified. This lectin was found to be relatively resistant to trypsin, and, particularly, to pepsin. All mice died within 48 hr when 200 microgram lectin per gram body weight was injected intraperitoneally and 14 microgram intravenously. The toxic activity changed in parallel with the lectin activity upon various treatments of the glycoprotein. In addition, blood analyses of injected mice suggested that the toxicity might be developed by the action of the lectin on blood cells.

Animals↗
PubMed1981

Quantitative studies on the polarization optical properties of living cells. I. Microphotometric birefringence detection system.

A method of polarization optical analysis is described in which phase retardation attributable to birefringence of a minute area in a microscopic object is determined. The optical system consists of a polarizing microscope with "rectified" strain-free lenses, a photoelectric detector to determine the intensity of the light passing through a minute window located at the image plane of the specimen, and a stage that moves the specimen at appropriate velocities for scanning. The error resulting from any flare of light emerging from outside of the area to be measured is minimized by limiting the illuminated area. The specimen can be observed during the measurement of light intensity by illuminating the whole microscope field at a wavelength different from that of the light used for the measurement. The retardation of the specimen is determined by comparing the specimen and background intensities as functions of the azimuth of a Brace-Köherl compensator. Alternatively, retardation is obtained directly from the light intensity at a fixed compensator angle, using the theory of polarization optics. The basal noise level for the present apparatus is approximately 0.03 nm when measuring birefringence of a 4-micron2 area in 0.1 s, using a X 40, NA 0.65 objective. The noise decreases in inverse proportion to the square root of the area times the duration of measurement.

Animals↗
PubMed1981

Quantitative studies on the polarization optical properties of living cells II. The role of microtubules in birefringence of the spindle of the sea urchin egg.

Birefringence of the mitotic apparatus (MA) and its change during mitosis in sea urchin eggs were quantitatively determined using the birefringence detection apparatus reported in the preceding paper (Hiramoto el al., 1981, J. Cell Biol. 89:115-120). The birefringence and the form of the MA are represented by five parameters: peak retardation (delta p), through retardation (delta t), interpolar distance (D1), the distance (D2) between chromosome groups moving toward poles, and the distance (D3) between two retardation peaks. Distributions of birefringence retardation and the coefficient of birefringence in the spindle were quantitatively determined in MAs isolated during metaphase and anaphase. The distribution of microtubules (MTs) contained in the spindle is attributable to the form birefringence caused by regularly arranged MTs. The distribution coincided fairly well with the distribution of MTs in isolated MAs determined by electron microscopy. Under the same assumption, the distribution of MTS in the spindle in living cells during mitosis was determined. The results show that the distribution of MTs and the total amount of polymerized tubulin (MTs) in the spindle change during mitosis, suggesting the assembly and disassembly of MTs as well as the dislocation of MTs during mitosis.

Animals↗
PubMed1980

An improved preparation of antibody-coated polystyrene beads for sandwich enzyme immunoassay.

An improved preparation of antibody-coated polystyrene beads for sandwich enzyme immunoassay of human thyroid-stimulating hormone (TSH) was described. Rabbit anti-TSH IgG was purified by eluting at pH 2.5 from a TSH-Sepharose column, diluted 3 or 9 fold with normal rabbit IgG and used for coating polystyrene beads by physical adsorption. In a sandwich enzyme immunoassay of TSH using rabbit (anti-TSH) Fab'-beta-D-galactosidase conjugate, beta-D-galactosidase activities specifically bound to thus prepared polystyrene beads in the presence of TSH was 2.8-6.3 fold higher than those bound to polystyrene beads coated with anti-TSH IgG before purification. A similar effect was observed when guinea pig anti-pork insulin IgG, rabbit (anti-human IgE) IgG and goat (anti-human IgE) IgG were treated at pH 2.5. This improvement may be based on a conformational change of Fc in IgG molecule which was caused by the treatment at pH 2.5. Other sandwich immunoassays such as fluoro- and radio-immunoassays may also be improved in the same way.

Animals↗
PubMed1979

An enzyme immunoassay for the measurement of thyroglobulin in human serum.

An enzyme-linked sandwich immunoassay using silicone rods coated with rabbit (anti-human thyroglobulin) immunoglobulin G and rabbit (anti-human thyroglobulin) monovalent fragment of immunoglobulin F (Fab') conjugated with beta-D-galactosidase was developed for the measurement of thyroglobulin in human serum. The volume of serum needed for the assay was as little as 2 microliters. The sensitivity of the assay was 3.5 ng/ml, which is equal to or rather higher than that of radioimmunoassay. The specificity of the assay was demonstrated by the following observations: (1) The absence of crossreaction of thyroxine and triiodothyronine, (2) non-detectability of thyroglobulin in the sera of patients who underwent total thyroidectomy, (3) parallelism of the standard curve with dilutions of reference serum. The precision of the assay was proven by the demonstration of the sufficient recovery of human thyroglobulin added to sera (92--99%) and coefficients of variance in within and between assays were 6.2--9.3 and 2.5--5.3%, respectively. Furthermore, a highly significant correlation was observed between thyroglobulin concentrations measured by our enzyme immunoassay and those by radioimmunoassay (r = 0.99, p less than 0.001, n = 63). Human thyroglobulin in serum was detectable in 90% of 146 normal subjects, the concentration (mean +/- S.D.) being 13.3 +/- 10.3 ng/ml.

Female↗
PubMed1979

Efficient conjugation of rabbit Fab' with beta-D-galactosidase from Escherichia coli.

An efficient procedure for the conjugation of rabbit Fab' with beta-D-galactosidase from Escherichia coli using N,N-o-phenylenedimaleimide is described. Thiol groups of Fab' were stabilized by the presence of ethylenediaminetetraacetate, and malemide groups were shown to be stable at pH 5 at 4 degrees C. The stability of thiol and maleimide groups enabled an efficient introduction of maleimide groups into Fab' and the average number of maleimide groups introduced into Fab' was 0.76 (range 0.73-0.79; n = 10) per molecule. As a result, 43.4% (range 41.3-46.9%; n = 6) of Fab' used could be conjugated with most of beta-D-galactosidase used. The average number of Fab' molecules conjugated per enzyme molecule was calculated to be 4.2 (range 4.0-4.6; n = 6). Both the enzyme and antibody activities were well preserved in the conjugate. There was no self-coupling of Fab', although the enzyme was polymerized to some extent during the conjugation reaction. The enzyme activity and cross-link in the conjugate was stable at pH 6.0-7.0 at 4 degrees C for at least 3 months.

Adsorption↗
PubMed1979

Improved procedure for the conjugation of rabbit IgG and Fab' antibodies with beta-D-galactosidase from Escherichia coli using N,N'-o-phenylenedimaleimide.

The procedures for the conjugation of rabbit IgG and Fab' antibodies with beta-D-galactosidase from Escherichia coli using N,N'-o-phenylenedimaleimide were improved in several respects as compared with the previous methods (Eur. J. Biochem. 62, 285--292, 1976; J. Immunol. 116, 1554--1560, 1976). Maleimide residues were efficiently introduced into antibodies under an atmosphere of nitrogen; the average number of maleimide residues introduced into IgG and Fab' antibodies were 0.78 (0.65--0.86) and 0.86 (0.80--0.95) per molecule, respectively. The conjugation with the enzyme was performed at 4 degrees C at pH 6.5 for 15 or more hours. The conjugates were almost completely separated from unreacted IgG and Fab' by gel filtration. When the recoveries of IgG, Fab', and beta-D-galactosidase in the conjugates were 23-29, 35-44, and 99%, respectively, the average numbers of IgG and Fab' molecules conjugated with the enzyme were 1.5-1.7 and 2.1-2.8 per molecule, respectively. There was no significant impairment of beta-D-galactosidase activity or the activity of anti-human IgG antibody to bind to human IgG upon conjugation. However, the conjugate preparation was heterogeneous, and one-third of each preparation consisted of aggregated conjugates less useful in sandwich enzymoimmunoassay than the remaining material. The conjugate with Fab' antibody gave lower control values in sandwich enzymoimmunoassay with silicone rubber as a solid phase than that with IgG antibody.

Animals↗