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Biomedical subjects

K Amako

Publications and source records attributed to K Amako.

At least 91 records · Page 5Linked to original sources
PubMed1987

Variability of haemolysin(s) produced by Vibrio vulnificus.

The peptide composition and antigenic cross-reactivity of partially purified and concentrated haemolysins of 16 strains of Vibrio vulnificus were examined by SDS-PAGE and immunoblotting analysis, using a monoclonal antibody (MAb), 6F8D, raised against the haemolysin. All strains produced a common peptide of 36 kDa and the MAb reacted with this peptide. In some strains, larger molecules, including a 56 kDa peptide, were produced, but the MAb did not react with this peptide. The haemolytic activity of the strains was effectively neutralized by the MAb, except in the case of strains producing the 56 kDa peptide. These findings indicate that the 36 kDa haemolysin is common to all 16 strains and that V. vulnificus can produce a second haemolysin which differs in molecular mass and antigenicity.

Antibodies, Monoclonal↗
PubMed1987

Monoclonal antibodies against the haemolysin of Vibrio vulnificus.

The extracellular haemolysin produced by Vibrio vulnificus strain FCC was partially purified from the culture supernate by sequential ammonium sulphate precipitation, gel filtration with Sepharose 4B, and DEAE-Sephacel ion-exchange column chromatography. Using this semi-purified haemolysin as the antigen, several monoclonal antibodies (MAbs) were established; they were all of the IgG2b class with lambda light chains. One representative MAb, 6F8D, completely neutralized the haemolytic activity and mouse lethal activity of extracellular toxin(s). In immunoblotting analysis of the peptides of the semi-purified haemolysin separated by SDS-PAGE, this MAb reacted, in particular, with a 36 kDa peptide. These findings suggest that the haemolysin is probably identical to the lethal toxin in the culture supernate of V. vulnificus strain FCC, which contained the 36 kDa peptide.

Antibodies, Monoclonal↗
PubMed1987

Protection against Campylobacter jejuni infection in suckling mice by anti-flagellar antibody.

We obtained two monoclonal antibodies of IgM class and IgA class of immunoglobulin prepared from mouse spleen cells immunized with crude flagellar preparation, and a polyclonal antibody raised against purified flagellin monomer of Campylobacter jejuni in a rabbit. The specificity of the reaction of these antibodies for flagellar filament was confirmed by Western blotting and by immunoelectron microscopy. These antibodies caused agglutination of the bacteria and inhibited the motility of the bacteria. When a strain of C. jejuni was treated with IgM class monoclonal antibody before being inoculated into suckling mice, it reduced colonization of the intestinal tract by this bacteria. Inhibition of the colonization by IgA class monoclonal antibody was less effective than that of IgM class, and the polyclonal antibody consisting mostly of IgG class immunoglobulin was without effect.

Animals↗
PubMed1987

Antigenic determinants on fimbriae of Serratia marcescens US5 analyzed using monoclonal antibodies.

The antigenic sites on small thin fimbriae of Serratia marcescens strain US5 were investigated using immunoelectron microscopy and monoclonal antibodies (MAbs). Negative staining of the fimbriae after treatment with MAbs showed a regularly spaced arrangement of the antibody molecules. When the subunit peptide was subjected to immunoblotting using the MAbs, a single band with a molecular weight of approximately 19kD was evident. This binding of the MAbs to the subunit peptide was completely abrogated after treatment with 2-mercaptoethanol, thereby suggesting the important role of disulfide linkage in the maintenance of the conformation of the antigenic site reacted with MAbs. Amino acid analysis of the subunit peptide revealed two cysteine residues, and cysteine residues were absent in the N-terminal portion.

Amino Acid Sequence↗
PubMed1987

Antigenic analysis of Serratia marcescens fimbriae with monoclonal antibodies.

Monoclonal antibodies (MAbs) were raised against the purified fimbriae of Serratia marcescens US46, a strain expressing three morphologically distinct fimbriae. The widths of these fimbriae were 7, 4.5, and 3 nm, respectively. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the purified fimbriae showed three bands with molecular weights of 21,000, 20,000, and 19,000, respectively. This strain had mannose-resistant (MR) hemagglutinating activity and was agglutinated by yeast cells. Therefore, strain US46 appeared to have both MR and mannose-sensitive fimbriae. In the immunoblot analysis, all MAbs reacted with the 20,000-molecular-weight subunit when given a choice of three differently sized subunits. Immunoelectron microscopy showed these MAbs attached to the MR fimbriae with the largest width (7 nm). The antigenic cross-reactivity of fimbriae was examined by an MAb-mediated agglutination test. All MR strains of S. marcescens and some mannose-sensitive strains were agglutinated by the MAbs. The serological homogeneity of MR fimbriae was confirmed by a spot test, using the crude purified fimbriae from several MR strains of S. marcescens. In other gram-negative rods, clinical isolates of Klebsiella spp. with hemagglutinating activity were agglutinated, but clinical isolates of Escherichia coli and Enterobacter spp. were not.

Antibodies, Bacterial↗
PubMed1987

Structure of the Staphylococcus aureus cell wall determined by the freeze-substitution method.

The fine structure of the Staphylococcus aureus cell wall was determined by electron microscopy with the new technique of rapid freezing and substitution fixation. The surface of the cell wall was covered with a fuzzy coat which consisted of fine fibers or an electron-dense mass. Morphological examination of the cell wall, which was treated sequentially with sodium dodecyl sulfate, trypsin, and trichloroacetic acid, revealed that this coat was partially removed by trypsin digestion and was completely removed by trichloroacetic acid extraction but was not affected by sodium dodecyl sulfate treatment, suggesting that the fuzzy coat consists mostly of a complex of teichoic acids and proteins. This was confirmed by the application of the concanavalin A-ferritin technique for teichoic acid and antiferritin immunoglobulin G technique for protein A.

Acetylglucosamine↗
PubMed1986

Structural changes in the nucleoid of Bacillus subtilis at low temperature.

The external shape of the nucleoid of Bacillus subtilis strain w23 was examined with a new electron microscopic technique, the rapid freezing and substitution fixation method. The nucleoid of the log and stationary phase cells was recognized as an area devoid of ribosomes and widely dispersed in the cytoplasm, which was different from that observed in OsO4-fixed cells. If the bacteria were exposed to low temperatures (0 to 10 C), the nucleoid showed a highly concentrated shape in the middle of the cytoplasm. These structural changes were observed only when the bacteria were maintained in a high-salt buffer. The results are discussed in relation to the membrane fluidity at low temperature.

Bacillus subtilis↗
PubMed1986

Astromicin-induced membrane damage in Serratia marcescens.

The morphological changes in Serratia marcescens induced by astromicin were determined by a new technique of electron microscopy, a rapid freezing and substitution fixation technique, and a freeze-fracturing technique. Two structural changes were observed. One was damage to the cytoplasmic membrane, and the other was the accumulation of a large electron-dense mass in the cytoplasm. The damage observed in the cytoplasmic membrane was the disappearance of the unit membrane structure from the thin-sectioned profile of the drug-treated bacteria and the loss of the membrane particles from the fractured surface of the membrane. Damage to the membrane was also suggested by the results of examination of the spheroplasts for stability. The spheroplasts prepared from the drug-treated bacteria were unstable in an osmotically controlled buffer. Most of the spheroplasts were lysed within 3 h, whereas those prepared from control cells were stable for more than 15 h. The electron-dense mass in the cytoplasm was usually seen in the polar region of the cell in close contact with the cell membrane. These structural changes were not specific for astromicin but were also found in gentamicin-treated cells.

Aminoglycosides↗
PubMed1986

Ontogeny of macrophage function to release superoxide anion in conventional and germfree mice.

To determine whether the presence of bacterial flora contributes to the ontogenic development of macrophage function, the ability of macrophages to release superoxide anion (O2-) in response to stimulation with phorbol myristate acetate was compared in conventional and germfree mice of various ages after birth. One-week-old conventional mice showed a very low level of O2- release by their macrophages, and gradual increases were observed in 2-, 3-, and 4-week-old mice in an age-dependent manner. Macrophages from germfree mice always showed a significantly lower level of O2- release compared with conventional mice of the same age; however, age-dependent functional development was seen also in the germfree group. The poor level of O2- release by macrophages from adult germfree mice could be restored to more than the level by conventional mice when the mice were conventionalized for 3 weeks. These results suggested that the ontogenic development of macrophage function is not controlled by the presence of bacterial flora but that the full-scale expression of function at each age is under the influence of microflora.

Age Factors↗
PubMed1986

Pulmonary defence mechanism in mice. A comparative role of alveolar macrophages and polymorphonuclear cells against infection with Candida albicans.

The protective roles of alveolar macrophages and polymorphonuclear cells were analyzed against intratracheal challenge with Candida albicans in mice. When mice were treated with carrageenan, a known cytotoxic agent for macrophages, there was no change in susceptibilities to the challenge in terms of the survival and the progressive elimination of fungi from the lung and kidney, in spite of a decreased in vitro phagocytosis of Candida albicans by their alveolar macrophages. On the other hand, irradiated mice (whole body irradiation with 800 rads) showed an enhanced mortality and a progressive growth of Candida albicans in their lungs and kidneys, although no change was observed in the in vitro phagocytic activity of alveolar macrophages until day 6 after irradiation. In normal and carrageenan treated mice, there was a progressive increase in the recruitment of polymorphonuclear cells into the lung after the challenge as shown by bronchoalveolar lavage and histological examination. In irradiated mice, on the other hand, there was a decreased recruitment of polymorphonuclear cells at 24 hr after the challenge, and a complete impairment at a late stage. When phagocytes were obtained from normal mice and examined for in vitro phagocytic activity to Candida albicans, polymorphonuclear cells showed higher activity than that of alveolar macrophages. These results suggest that polymorphonuclear cells play a very important role in the protection against intratracheal infection with Candida albicans.

Animals↗
PubMed1986

Enhanced clearance of Candida albicans from lung after intratracheal immunization.

The comparative effect of immunization by intratracheal and intraperitoneal routes was assessed against intratracheal challenge with Candida albicans. When mice were immunized intratracheally with 5 X 10(6) cfu of viable or heat-killed C. albicans 2 weeks before challenge, they showed an enhanced resistance against intratracheal challenge with C. albicans, in spite of a decrease in phagocytosis of C. albicans in vitro by alveolar macrophages from intratracheally immunized mice. In these mice, 100% survival and an accelerated elimination of C. albicans from their lungs were observed after intratracheal challenge with C. albicans. In contrast, intratracheal immunization with same high dose of viable or heat-killed C. albicans 1 week before challenge, and intraperitoneal immunization with same high or low doses of viable or heat-killed C. albicans 1 or 2 weeks before intratracheal challenge with C. albicans, could not induce such high resistance against intratracheal challenge with same high dose of viable C. albicans. Comparing to normal and intraperitoneally immunized mice, number of C. albicans recovered from the lung 48 hr after challenge was approximately 100 fold smaller in mice immunized by intratracheal route 2 weeks before challenge. When cellular response was determined by bronchoalveolar lavage after intratracheal inoculation of C. albicans, a prominent recruitment of polymorphonuclear cells was observed 12 and 36 hr after challenge in control mice, intraperitoneally and intratracheally immunized mice. Among these groups, sustained polymorphonuclear cell recruitment by 72 hr after challenge was seen only in the group immunized by intratracheal route with high dose of heat-killed C. albicans 2 weeks before challenge.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗