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At least 739 records · Page 41Linked to original sources
PubMed1990

Reactivity of Limulus amoebocyte lysate towards (1----3)-beta-D-glucans.

The structure activity relationship for beta-D-glucans for the gelation of the amoebocyte lysates of the horseshoe crab (Limulus) has been investigated. beta-D-Glucans that had no (1----3) linkages induced little or no gelation. The (1----3)-beta-D-glucans curdlan (unbranched), grifolan (approximately 33% branched), schizophyllan (approximately 33% branched), lentinan (approximately 40% branched). SSG (approximately 50% branched), and OL-2 (approximately 66% branched) induced significant gelation. The optimum concentration for gelation was correlated with the content of branching. Single chain (rather than a triple helix) conformation and higher molecular weight were associated with higher reactivity.

Animals↗
PubMed1991

Relationship between conformation and biological response for (1----3)-beta-D-glucans in the activation of coagulation factor G from limulus amebocyte lysate and host-mediated antitumor activity. Demonstration of single-helix conformation as a stimulant.

The relationship between the conformation of (1----3)-beta-D-glucans in gel or hydrated form and the stimulation of two types of biological responses, namely, activation of coagulation Factor G from limulus amebocyte lysate (LAL) and host-mediated antitumor activity was examined. Both types were activated by the single-helical conformation, as revealed by high-resolution, solid-state 13C-n.m.r. spectroscopy. The potency of activation of Factor G was increased over 100-fold by treatment with a NaOH solution which leads to a complete or partial conversion from the triple to the single helix. Such a single-helix specific response was also demonstrated for the antitumor activity of curdlan, although the distinction was less pronounced for branched (1----3)-beta-D-glucans. The presence of the single-helix conformation was observed in schizophyllan gel, even though the triple helix is the most stable form of branched glucans in aqueous media.

Animals↗
PubMed1996

Evaluation of endotoxiuria for diagnosis of urinary tract infection after major surgical procedures.

In 148 patients after major surgical procedures urinary endotoxin levels were determined and compared with bacteriological results. The study was designed as a screening study. Urine samples were collected once by suprapubic or transurethral catheters. In a first series of 49 patients urine bacteriology was positive (mainly, Gram-negative rods were found) in 3 cases. However, endotoxin determination was positive in these 3 patients and in a further 10 patients receiving antibiotic therapy for other reasons. Therefore, the following 99 patients were studied also by urinalysis by reagent strips for leukocytes and nitrite. In the second series, 12 urine cultures positive for bacteria were observed. Eleven samples were also endotoxin positive. Five more patients were endotoxin positive and had pathological but unspecific reagent strip results. These patients were treated with antibiotics for other reasons. Patients with candida found in the urine culture (n = 5) were endotoxin negative. Thus, endotoxin determination in urine obtained by suprapubic or transurethral catheters proved to be a very sensitive method for diagnosis of bacterial contamination, even during antibiotic treatment.

Adult↗
PubMed1978

A rapid, novel method for the solid-phase derivatization of IgG antibodies for immune-affinity chromatography.

We show that a solid-phase immune adsorbent can be prepared from rabbit antiserum by isolating the IgG fraction with the A protein of Staphylococcus aureus associated covalently with a Sepharose matrix. The IgG is then coupled to the matrix using the cross-linking agent dimethylsuberimidate. IgG antibody bound in this fashion is in the proper orientation for combination with antigen because association with protein A occurs via the Fc portion of the IgG molecule, thus leaving the combining site of the molecule free to interact with antigen.

Antibodies, Anti-Idiotypic↗
PubMed1982

Elicitation of peritoneal polymorphonuclear neutrophils from mice.

Although the mouse has been used extensively as a model for the study of host-parasite relationships, murine neutrophils have not been used nearly as often as PMNs from other species for in vitro functional assays due to lack of a commonly used procedure for murine neutrophil collection. These studies compared two eliciting agents and characterized the phagocytic and bactericidal activity of murine polymorphonuclear neutrophils elicited from the peritoneal cavity. We examined the effects of mouse strain (BALB/c, C57BL/6 and DBA/2) and sex, eliciting agent (0.2% glycogen vs. 3% fluid thioglycolate medium) and donor sacrifice method (ether vs. cervical dislocation) on the number of neutrophils recovered in peritoneal exudate. The greatest number of neutrophils was harvested when mice were sacrificed 5 h after intraperitoneal injection of 2.5 ml of 3% thioglycolate medium. This method as described allows reproducible collection of adequate numbers of neutrophils for use in in vitro assays of neutrophils phagocytic and bactericidal function.

Animals↗
PubMed1984

Evaluation of absorption with Limulus amebocyte lysate to remove contaminating endotoxin from interferon and lymphokine preparations.

Alpha and beta human interferon (IFN) preparations and lymphokines (supernatants of PHA-stimulated blood lymphocytes) were deliberately contaminated with endotoxin (20 ng/ml) and subsequently rendered endotoxin-free by absorption with Limulus amebocyte lysate (LAL). Absorption with LAL did not appreciably affect the antiviral activity of IFN and lymphokines in 8 experiments and caused a 30-50% reduction in two. The capacity of these agents to stimulate natural killer cell activity and monocyte cytotoxicity was not consistently modified by absorption on LAL. When the chemotactic activity of lymphokine for monocytes was measured, the maximal number of monocytes induced to migrate and the maximal active lymphokine concentration were not affected by absorption with LAL. LAL-treated lymphokines, however, showed a prozone phenomenon, presumably related to the release of chemotaxis inhibitor(s) from the LAL gel.

Absorption↗
PubMed1990

Removal of endotoxin from protein solutions by phase separation using Triton X-114.

Endotoxin contamination of protein solutions was reduced by a phase separation technique using the detergent, Triton X-114. Protein solutions containing endotoxin were treated with Triton X-114 on ice. The solution was then warmed to 37 degrees C, whereupon two phases formed. The Triton X-114 phase, containing the endotoxin, was precipitated by centrifugation. The first cycle of phase separation produced a 1000-fold reduction of endotoxin from contaminated preparations of cytochrome c, catalase and albumin. Complete removal of endotoxin could be achieved by further cycles of phase separation. Each cycle of phase separation resulted in only a 2% loss of protein, and could be completed within 15 min. The small amount of detergent (0.018%) that persisted in protein solution could be removed by gel filtration or absorption. Proteins treated by this procedure retained normal functions. This phase separation technique provides a rapid and gentle method for removing endotoxin from protein solutions.

Catalase↗
PubMed1990

Cross-reactive affinity purification of immunoglobulin recognizing common gram-negative bacterial core antigens.

A procedure isolating immunoglobulins specific for common gram-negative bacterial core antigens is described. A polyclonal reagent was purified by ammonium sulfate precipitation, dialysis, and column affinity chromatography. The initial vaccinal antigen was an Ra mutant Escherichia coli O111:B4 (strain J5). The capture antigen was lipopolysaccharide derived from an Ra mutant, Salmonella typhimurium TV119 covalently-linked to an agarose matrix. Column eluants were characterized in terms of total protein concentration, IgG concentration, and EIA titer recognizing E. coli (J5). Low protein, low IgG, high EIA reading fractions were isolated, demonstrating the utility of the described technique to purify broad spectrum cross-reactive immunoglobulin reagents.

Animals↗
PubMed1994

Semi-preparative purification and validation of monoclonal antibodies for immunotherapy in mice.

A number of rat hybridomas were adapted to grow in RPMI containing either 5% IgG-depleted FCS or 1% serum-free Nutridoma. Alternatively, protein-free Ultradoma PF was used. Growth in these media allowed purification procedures to be used that are based on tangential ultrafiltration in combination with affinity chromatography on gels linked to protein G or anti-rat L chain coupled antibodies. The isolated antibody preparations were found to be pure and to consist of monomeric intact IgG. The yield and recovery of mAb using this procedure were found to be consistently high. These antibody preparations were analyzed for endotoxin contamination. Whereas during isolation endotoxin contamination increased, the endotoxin content per mg purified protein did not. Affinity chromatography on Detoxi-gel resulted in the efficient removal of this contamination and using this protocol the antibody preparations obtained were found to be of sufficient purity, activity and low endotoxin content to permit their in vivo use in animal models of immunotherapy.

Animals↗
PubMed1988

A glycolipid precursor of bacterial lipopolysaccharide (lipid X) lacks activity against endothelial cells in vitro and is not toxic in vivo.

Lipid X (2,3-diacylglucosamine-1-phosphate) accumulates in mutants of Escherichia coli incapable of assembling the disaccharide backbone of lipid A, the principle endotoxic moiety of bacterial lipopolysaccharide (LPS). We compared the effects of lipid X on cultured bovine aortic endothelial cell (BEC) viability and prostacyclin (PGI2) release with those of lipid A and LPS. At 10(-5) M, both LPS and lipid A produced significant BEC cytotoxicity (percentage cytotoxicity 69 +/- 4 for LPS and 51 +/- 11 for lipid A) and induced a variable but consistent increase in the release of PGI2 (11- to 73-fold increase for LPS and 4- to 6-fold increase for lipid A). Lipid X, in contrast, was not toxic and did not induce PGI2 release at 10(-4) M. Pretreatment and coincubation of BEC with lipid X, at a concentration 100 times greater than LPS, failed to prevent LPS-mediated cytotoxicity. Intravenous infusion of lipid X in goats had no effect except for a modest elevation in the pulmonary artery pressure during the period of infusion. Moreover, pretreatment of goats with lipid X (70 micrograms/kg) did not block the effects of a subsequent infusion of LPS (5 micrograms/kg). These data suggest that a fatty acid-substituted disaccharide is the minimal molecular requirement for the numerous effects in vivo and activity in vitro induced by LPS. Furthermore, these effects are not prevented by pretreatment with a monosaccharide precursor of lipopolysaccharide, lipid X, at a dose 10- to 100-fold greater than that of LPS.

6-Ketoprostaglandin F1 alpha↗
PubMed1982

Endotoxic activity in periapical lesions.

Thirty tissue samples were examined histologically and classified as being inflamed (apical granulomas) or noninflamed (scars or noninflamed cysts). The samples were then homogenized in pyrogen-free water and treated to remove interfering substances. The presence of endotoxin was then determined by means of the limulus assay; 75 percent of the inflamed tissues were positive for endotoxin, while only 20 percent of the noninflamed tissues contained endotoxin. The presence of endotoxin was thus highly correlated (p = 0.015) with the presence of inflammation in these tissues.

Endotoxins↗
PubMed1994

Endotoxin-induced platelet aggregation in heparinised equine whole blood in vitro.

Endotoxaemia is a leading cause of death among horses. Thrombocytopenia is a common finding in clinical and experimentally-induced cases of endotoxaemia and can lead to coagulopathies, including disseminated intravascular coagulopathy which is usually fatal. In this study it was shown that endotoxin (3 ng ml-1 to 25 micrograms ml-1) can aggregate equine platelets in heparinised whole blood in vitro. The endotoxin-induced aggregation (EIA) was shown to be dependent on the presence of leucocytes in the blood and did not occur when detoxified endotoxin was used, suggesting that lipid A was necessary for the response. Aspirin (1 mmol litre-1) had no effect on EIA whereas apyrase (40 micrograms ml-1) completely abolished it and CV3988 (3 to 30 mumol litre-1) (a competitive antagonist of platelet-activating factor) inhibited the response in a concentration-dependent manner. It is concluded that endotoxin activates equine platelets at low concentrations through an indirect mechanism that involves calcium, leucocytes, adenine nucleotides and platelet-activating factor.

Animals↗
PubMed1988

Endotoxaemia in complicated falciparum malaria.

In a prospective hospital-based study, endotoxin was detected by amoebocyte limulus lysate test in the blood of 18 of 20 patients with complicated Plasmodium falciparum (16 with cerebral malaria, 2 with blackwater fever, one with acute malarial hepatitis and one with hepatorenal failure) and in all 5 patients with uncomplicated malaria tested, but in none of 5 healthy volunteers. There were 4 deaths among the 18 patients with complicated malaria and endotoxaemia. No correlation between endotoxaemia and presence of complications, clinical severity, or degree of parasitaemia was found. A concomitant bacterial infection could account for endotoxaemia in 11 of the 16 patients with cerebral malaria and endotoxaemia; in the other 5 patients with cerebral malaria, 4 with other complications, and 5 with uncomplicated malaria, endotoxin was detected in the blood without any evidence of bacterial infection.

Adolescent↗