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S Kozaki

Publications and source records attributed to S Kozaki.

At least 91 records · Page 5Linked to original sources
PubMed1988

Production of monoclonal antibody against Aeromonas hydrophila haemolysin.

Two hybridoma cell lines that produce monoclonal antibodies against Aeromonas hydrophila haemolysin were established by fusion of myeloma and spleen cells obtained from a mouse immunised with haemolysin detoxified with tetranitromethane. Enzyme-linked immunosorbent assay (ELISA) showed that the two purified monoclonal antibodies, B7 and B11, recognised the same epitope on the haemolysin molecule. Antibody B7 neutralised the haemolytic and enterotoxic activities of the haemolysin. It is concluded that the same site on the haemolysin molecule is responsible for both haemolytic and enterotoxic activities.

Aeromonas↗
PubMed1987

Development of anti-erythrocyte antibodies in mice infected with Babesia rodhaini.

Anti-erythrocyte and anti-parasite antibodies were detected in sera of mice infected with a lethal strain of Babesia rodhaini by an enzyme linked immunosorbent assay (ELISA). Both antibodies were produced soon after infection, and their levels increased rapidly in parallel with increasing parasitemias. The number of erythrocytes decreased as parasitemias increased. Serum levels of both antibodies increased in mice injected with sonicated parasite antigens derived from infected erythrocytes, but not in mice injected with a similar preparation derived from non-infected erythrocytes. Immunoglobulins were detected by on ELISA on the membrane of erythrocytes from mice infected with B. rodhaini or injected with the sonicated antigens. The erythrocytes-associated immunoglobulins seemed to be mainly IgG. These results indicate that B. rodhaini infections induce the production of anti-erythrocyte antibodies as well as anti-parasite antibodies, and the parasite antigens may contain some factors capable of inducing the anti-erythrocyte autoimmune responses.

Animals↗
PubMed1987

Antigenic structure of Clostridium botulinum type B neurotoxin and its interaction with gangliosides, cerebroside, and free fatty acids.

A fragment distinct from the heavy and light chains was obtained by treatment of Clostridium botulinum type B neurotoxin with chymotrypsin. Enzyme-linked immunosorbent assay and immunoblotting analysis with monoclonal antibodies showed that the fragment consisted of the light chain and part of the heavy chain (H-1 fragment) linked together by a disulfide bond. Monoclonal antibodies reacting to the heavy chain but not to the fragment were thought to recognize the epitopes on the remaining portion (H-2 fragment) of the heavy chain, being easily digested by chymotrypsin. Thus, the antigenic structure of type B neurotoxin resembles those of type A and E neurotoxins. The chymotrypsin-induced fragment bound to cerebroside and free fatty acids but not to gangliosides. The manner of binding of type B neurotoxin to gangliosides and free fatty acids was different from those of type A and E neurotoxins. Such differences in the reactivities to lipids may be related to the finding that each neurotoxin binds to a type-specific site on the neural membrane.

Antibodies, Monoclonal↗
PubMed1987

Activities of Aeromonas hydrophila hemolysins and their interaction with erythrocyte membranes.

The activity of hemolysin produced by Aeromonas hydrophila CA-11, isolated from an environmental source, was more sensitive to temperature than that of hemolysin produced by strain AH-1, isolated from a diarrheal case. CA-11 hemolysin failed to elicit hemolysis below 10 degrees C. Immunoblotting analyses showed that both hemolysins formed into oligomers in rabbit erythrocyte membrane even when no hemolysis occurred. suggesting that the binding and the subsequent oligomerization are temperature independent. Sodium salicylate inhibited lysis of rabbit erythrocytes by both hemolysins, but selected monosaccharides and oligosaccharides did not. Thin-layer immunostaining indicated that both hemolysins bound to phosphoglycerides with net negative charge but weakly to the ones with no net negative charge. Neither sphingomyelin nor lysophosphoglyceride reacted with the hemolysins, whereas the hemolysins bound to free fatty acids. These results suggest that the binding of either hemolysin to the membrane component, probably phospholipid, requires both negative charge of the polar head group and suitable hydrophobicity of the nonpolar tails.

Aeromonas↗
PubMed1987

Characterization and localization of Paragonimus westermani antigen stimulating antibody formation in both the infected cat and rat.

Applicability of the adult Paragonimus westermani antigen for detection of anti-immature P. westermani antibodies in experimentally infected rats, a paratenic host of this lung fluke, was examined. The serum antibodies of the cats and rats infected with P. westermani metacercariae were detected by enzyme-linked immunosorbent assay (ELISA) with the adult-fluke antigen. The ELISA titers of serum samples of the rats infected with only immature flukes were as high as those of the cats infected with adult flukes. Sodium dodecyl sulfate polyacrylamide gel electrophoresis and the immunoblotting technique showed that a major protein band of 27,000 daltons was recognized in the sera of the infected cats and rats. Immunoperoxidase staining applied on the sectioned flukes provided evidence showing that the antigenic substance was located on the surface of the gut epithelium and in the luminal contents in both adult and immature flukes. The adult-fluke antigen containing the 27,000-dalton substance is applicable as a standard antigen for diagnosis of paragonimiasis westermani in not only definitive hosts but also in paratenic hosts.

Animals↗
PubMed1986

Evidence for direct binding of Clostridium botulinum type E derivative toxin and its fragments to gangliosides and free fatty acids.

Clostridium botulinum type E derivative toxin and its heavy chain bound to gangliosides GT1b, GD1a and GQ1b and saturated and unsaturated free fatty acids with chain lengths of 14-20 carbons. The L-H-1 fragment lacking the carboxyl-terminal portion of the heavy chain bound to free fatty acids but not to gangliosides. These observations led us to a new hypothesis on the mechanism of binding between botulinum toxin and gangliosides; the carboxyl-terminal portion (H-2 fragment) of the heavy chain binds to an oligosaccharide residue of gangliosides and then the amino-terminal portion (H-1 fragment) interacts with the hydrophobic portion of gangliosides consisting of fatty acids.

Binding Sites↗
PubMed1986

Effects of Ca2+ and other cations on the action of Clostridium perfringens enterotoxin.

We investigated the role of extracellular Ca2+ in the Clostridium perfringens enterotoxin-induced alteration of the permeability of the plasma membrane. Enterotoxin released 86Rb and 51Cr from the Vero cells preloaded with the isotope. In the presence of EGTA, however, it released 86Rb but not 51Cr. The binding of enterotoxin to the cells was not influenced by Ca2+ or Mg2+. The effects of various cations on the enterotoxin-induced 51Cr release was also studied. The release depended on extracellular Ca2+ but not on Mg2+; it was inhibited by each of Zn2+, La3+ and Co2+. Zn2+ and Co2+ also inhibited 51Cr release caused by the enterotoxin previously bound to the cell membrane. In contrast, antibody against enterotoxin did not neutralize the toxin once it was bound to the Vero cells. When the cells were treated with enterotoxin, 45Ca influx occurred and reached the plateau in a few minutes, as did 86Rb release.

Animals↗
PubMed1986

TLC immunostaining characterization of Clostridium botulinum type A neurotoxin binding to gangliosides and free fatty acids.

The receptor structure of Clostridium botulinum neurotoxin type A was analysed by TLC immunostaining. GQ1b was found to be the most potent receptor, and the neurotoxin also bound to GT1b and GD1a, but not to GM3, GM2, GM1, GD3, GD1b and GT1a. Optimum binding of neurotoxin to the ganglioside appeared in 0.01 M phosphate buffer (pH 7.2) containing 0.2% NaCl. Higher and lower NaCl concentrations diminished neurotoxin binding to the ganglioside. In addition, the neurotoxin was able to bind to free fatty acids. Maximum binding was observed on stearic acid and neurotoxin binding to free fatty acids was not affected by NaCl concentration.

Botulinum Toxins↗
PubMed1986

Production and characterization of monoclonal antibodies to Clostridium perfringens enterotoxin.

Four hybridoma cell lines producing monoclonal antibodies to Clostridium perfringens enterotoxin were established by fusion of mouse myeloma and spleen cells obtained from mice immunized with the enterotoxin and its toxoid. An enzyme-linked immunosorbent assay indicated that the two antibodies, 2-B-4 and 3-G-10, bound to those regions that were located close each other; the others, 3-B-2 and 2-H-2, bound to other independent regions on the enterotoxin. Release of 51Cr from Vero cells with the enterotoxin was inhibited by either 2-B-4 or 3-G-10, both of which inhibited the binding of 125I-labeled enterotoxin to the cells. Neither binding nor cytotoxicity of the enterotoxin was affected by 2-H-2; 3-B-2 only barely inhibited the binding but neutralized the enterotoxin shown by 51Cr release. It seems justified to conclude that 3-B-2 blocks the toxic action after the enterotoxin has bound to Vero cells.

Animals↗
PubMed1986

The use of monoclonal antibodies to analyze the structure of Clostridium botulinum type E derivative toxin.

Six monoclonal antibodies against Clostridium botulinum type E derivative toxin were prepared. Three of the five binding to the heavy chain neutralized the derivative toxin; the other one binding to the light chain did not. Immunoblotting analysis with the monoclonal antibodies showed that the fragment obtained by tryptic digestion consisted of the light chain and part of the heavy chain (H-1 fragment) linked together by a disulfide bond(s) and that the antigenic determinants common between type E and F derivative toxins were located on both the heavy and light chains. The fragment induced by chymotrypsin treatment, like the tryptic fragment, bound to four monoclonal antibodies. The mild tryptic treatment and reduction resulted in separation of the chymotryptic fragment into two smaller fragments corresponding to the light chain and H-1 fragment. These results indicate that H-1 fragment contains the amino-terminal portion of the heavy chain. The monoclonal antibody neutralizing the toxin and probably recognizing the epitope on the carboxyl-terminal portion (H-2 fragment) of the heavy chain effectively competed for binding of 125I-labeled derivative toxin to synaptosomes. Of the two monoclonal antibodies neutralizing the toxin and recognizing the epitopes on H-1 fragment, one partially inhibited binding, but the other did not. This suggests that the binding of 125I-labeled derivative toxin depends mainly on the carboxyl-terminal region of the heavy chain and that interference with binding is not the only means of toxin neutralization.

Animals↗
PubMed1986

Production of "Asao toxin" by Aeromonas strains isolated from feces and drinking water.

Cultures of Aeromonas species were tested for production of a toxin recently purified by Asao et al. (T. Asao, Y. Kinoshita, S. Kozaki, T. Uemura, and G. Sukaguchi, Infect. Immun. 46:122-127, 1984) and described as a hemolysin with enterotoxic and cytotoxic activity. The toxin was produced by only 63% of Aeromonas sobria strains and by 93% of Aeromonas hydrophila strains. Also, 54% of A. hydrophila strains produced another cytotoxic entity.

Aeromonas↗
PubMed1986

Purification and characterization of an Aeromonas hydrophila hemolysin.

A hemolysin produced by Aeromonas hydrophila CA-11, isolated from an environmental source, was purified by sulfopropyl-Sephadex C-25 chromatography at pH 5.0. This hemolysin caused fluid accumulation in infant mouse intestines and rabbit intestinal loops and killed Vero cells, as did the hemolysin produced by strain AH-1, isolated from a diarrheal case. In polyacrylamide gel electrophoreses at pHs 4.0 and 9.4 and in thin-layer isoelectric focusing, CA-11 hemolysin migrated as a single band to a position different from that of AH-1 hemolysin. Immunodiffusion tests indicated that CA-11 hemolysin was immunologically related to AH-1 hemolysin but possessed unique antigenic determinants. Neutralization tests with antihemolysin sera also demonstrated immunological cross-reactivity between AH-1 and CA-11 hemolysins. These results apparently indicate that the hemolysins produced by the two strains of A. hydrophila are immunologically and physicochemically different from each other.

Aeromonas↗
PubMed1984

The role of sialic acid in the nerve terminal for the release of transmitter.

The role of sialic acid in the frequency of miniature endplate potentials (MEPPs) was examined using neuraminidase and gangliosides in the mouse diaphragm. Neuraminidase increased and decreased MEPP frequency in normal K+ and high K+ solution, respectively. The effects were dependent on the presence of Ca2+ in extracellular medium. Neuraminidase liberated sialic acid from and lowered Ca2+-binding capacity of synaptosomal membrane. Gangliosides treatment of the tissue partially restored the effects of neuraminidase on the frequency of MEPP and Ca2+-binding capacity. It is possible that sialic acid in the nerve endings provides a functional storage site which supply intracellular Ca2+ to cause a transmitter release.

Animals↗
PubMed1984

Purification and some properties of Aeromonas hydrophila hemolysin.

A hemolysin produced by a strain of Aeromonas hydrophila isolated from a patient with diarrhea was purified by acid precipitation and quarternary aminoethyl-Sephadex chromatography. The molecular weight of the hemolysin was estimated at 50,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and at 48,000 by Sephadex G-100 gel filtration. In polyacrylamide gel electrophoresis at pH 4.0 and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the hemolysin migrated as a single band, whereas electrophoresis at pH 9.4 and thin-layer isoelectric focusing demonstrated multiple bands. The results may indicate charge isomers of the hemolysin. The purified hemolysin had a hemolytic activity of 134 hemolytic units per microgram of protein on rabbit erythrocytes. It caused fluid accumulation in infant mouse intestines and rabbit ligated ileal loops. Purified hemolysin also elicited cytotoxicity to Vero cells and lethal toxicity to mice. All these biological activities were lost on heating for 5 min at 56 degrees C. These findings support the notion that A. hydrophila hemolysin is a cytotoxic enterotoxin.

Aeromonas↗