Biomedical subjects

F S Chu

Publications and source records attributed to F S Chu.

At least 55 records · Page 3Linked to original sources

PubMed1987

Production and characterization of a generic antibody against group A trichothecenes.

An antibody against group A trichothecenes was produced after immunization of rabbits with an immunogen prepared by conjugation of T-2 toxin to bovine albumin at the C-8 position. T-2 toxin was first converted to 3-acetylneosolaniol (3-Ac-NEOS) and then to its hemisuccinate (HS) before conjugation to the protein. The rabbits showed a quick immune response after immunization of the new conjugate. The antibody produced bound with tritiated T-2 toxin, T-2 tetraol tetraacetate, and diacetoxyscirpenol (DAS) and showed good cross-reactivities with most of the group A trichothecenes. The concentrations causing 50% inhibition of binding of 3H-T-2 toxin to the new antibody by unlabeled T-2, acetyl-T-2, 3'-OH-T-2, DAS, 3-Ac-NEOS-HS, 3'-OH-Ac-T-2, T-2 tetraol tetraacetate, iso-T-2, 3-Ac-NEOS, Ac-DAS, and 3,4,15-triacetyl-7-deoxynivalenol were found to be 0.34, 0.34, 0.6, 2.5, 4, 10, 18, 24, 100, 200, and 300 ng/assay, respectively; for HT-2, T-2 triol, and T-2 tetraol, the concentration was greater than 1000 ng/assay. Nivalenol, deoxynivalenol (DON), 15-acetyl-DON, and triacetyl-DON, did not inhibit the binding at 1000 ng/assay. The practical application of using this new antibody for radioimmunoassay (RIA) of trichothecene was tested by spiking T-2 toxin to corn. T-2 toxin was then extracted with acetone, subjected to a simple Sep-Pak C-18 reversed-phase treatment, and analyzed by RIA. The overall recovery for 18 samples spiked with 10 to 50 ppb of T-2 toxin was 94.22%.

Animals↗

PubMed1987

Production and characterization of antibodies against HT-2 toxin and T-2 tetraol tetraacetate.

Three new immunogens which were prepared by conjugation of the carboxymethyl oxime (CMO) derivatives of HT-2 toxin, T-2 tetraol (T-2 4ol), and T-2 tetraol tetraacetate (T-2 4Ac) to bovine serum albumin (BSA) were tested for the production of antibodies against the major metabolites of T-2 toxin. Antibodies against HT-2 toxin and T-2 4Ac were obtained from rabbits 5 to 10 weeks after immunizing the animals with CMO-HT-2-BSA and CMO-T-2 4Ac-BSA conjugates. Immunization with CMO-T-2 4ol-BSA resulted in no antibody against T-2 4ol. The antibody produced against HT-2 toxin had great affinity for HT-2 toxin as well as good cross-reactivity with T-2 toxin. The relative cross-reactivities of anti-HT-2 toxin antibody with HT-2 toxin, T-2 toxin, iso-T-2 toxin, acetyl-T-2 toxin, 3'-OH HT-2, 3'-OH T-2, T-2 triol, and 3'-OH acetyl-T-2, were 100, 25, 10, 3.3, 0.25, 0.15, 0.12 and 0.08%, respectively. Antibody against CMO-T-2 4Ac was very specific for T-2 4Ac and had less than 0.1% cross-reactivity with T-2 toxin, HT-2 toxin, acetyl-T-2 toxin, diacetoxyscirpenol, deoxynivalenol, and deoxynivalenol triacetate as compared with T-2 4Ac. The detection limits for HT-2 toxin and T-2 4ol by radioimmunoassay were approximately 0.1 and 0.5 ng per assay, respectively.

Animals↗

PubMed1986

Improved method for production of antibodies against T-2 toxin and diacetoxyscirpenol in rabbits.

A new, improved approach for the production of antibodies against T-2 toxin and diacetoxyscirpenol (DAS) was developed. The method involves the use of immunogens which were prepared by conjugating O-carboxymethoxyl oxime (CMO) derivatives of both toxins to bovine serum albumin (BSA). Isomers a and b of CMO-T-2 toxin and isomer b of CMO-DAS were tested. Antibodies against both toxins were demonstrated as early as 4 weeks after immunization. a-CMO-T-2-BSA conjugate was a better immunogen than the b isomer, and the highest titers (6,000) were reached 14 weeks after immunization and one booster injection. Antibody titers for rabbits immunized with the b isomer of CMO-T-2 never reached more than 2,000. The specificity of antibodies obtained from rabbits after immunization with CMO-T-2-BSA was similar to that of hemisuccinate-T-2-BSA. Anti-b-T-2 antibodies had slightly higher cross-reactivity with H-T-2 toxin than did the antibody obtained from rabbits immunized with the conjugate of the a isomer. The relative cross-reactivities of anti-a-CMO-T-2 antibody with T-2, acetyl-T-2, H-T-2, T-2-triol, 3'-OH-T-2, and T-2 tetraol were 1, 4.5, 5.7, 250, 500, and 3,000, respectively. The relative cross-reactivities of anti-b-T-2 antibody with T-2, acetyl-T-2, H-T-2, and T-2 triol were 1, 2, 3, and 488, respectively. Antibodies against b-CMO-DAS showed a high degree of cross-reactivity with monoacetoxyscirpenols (MAS). The relative cross-reactivities of anti-B-DAS antibody with DAS, 4-MAS, 15-MAS, acetyl-deoxynivalenol, T-2-toxin, acetyl-T-2, and neosolaniol were 1, 4, 5, 76, 107, 147, and 266, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1985

Modification of in vitro metabolism of T-2 toxin by esterase inhibitors.

In vitro metabolism of T-2 toxin with S-9 fraction obtained from livers of phenobarbital-treated pigs and rats in the presence of different esterase inhibitors, including NaF, p-hydroxymercuribenzoate, phenylmethylsulfonyl fluoride, eserine sulfate, diisopropylfluorophosphate, and diethyl p-nitrophenyl phosphate, was studied. The metabolism was completely shifted to the hydroxylation at the C-3' position in the T-2 toxin molecule when esterase inhibitors were present. Diethyl p-nitrophenyl phosphate was found to be the most potent among six esterase inhibitors tested. In the presence of 10(-4) M diethyl p-nitrophenyl phosphate, 3'-hydroxy-T-2 toxin was the only metabolite detected. Similar results were obtained when other T-2-related metabolites were tested. The yield of conversion of T-2 toxin, acetyl T-2 toxin, HT-2 toxin and T-2 triol to their respective 3'-hydroxyl derivatives were 82, 73, 72, and 75%, respectively.

Animals↗

PubMed1984

Immunohistochemical fate of ochratoxin A in mice.

By a highly specific antibody against ochratoxin A (OA), the immunohistochemical fate of po administered OA in Swiss mice given a single dose of 25 mg/kg was examined by light microscopy with two immunoperoxidase methods, an indirect method and a peroxidase-antiperoxidase method. Immunohistochemical staining for OA was demonstrated in the esophagus, stomach, small intestine, kidneys, and liver. OA was most intense in the gastrointestinal tract, intermediate in the kidneys, and least in the liver. In the gastrointestinal tract, OA was demonstrable: (i) in the squamous layer of the esophagus, (ii) in the surface mucus and within macrophages and neutrophils of the stomach lamina propria, (iii) within the cytoplasm of duodenal and jejunal surface epithelial cells as well as within macrophages and neutrophils of the duodenal and jejunal lamina propria. Maximal gastrointestinal tract staining was evident from 5 min to 3 hr, while the esophagus remained stained for 24 hr postdosing. No evidence of OA was obtained in the ileum. In the kidneys, the vast majority of OA was localized within the cytoplasm of the proximal convoluted tubular cells as brown-stained vacuoles, whose size became largest at 3 hr postdosing. Staining for OA was also demonstrated within the epithelium of distal convoluted tubules, the macula densa, the loop of Henle, and sometimes within the epithelium of Bowman's capsule and glomerulus. OA in hepatocytes was most intense between 40 min and 3 hr postdosing. Stained hepatocytes were slightly more concentrated in the periportal area than in the area of the central vein and typically exhibited strong cytoplasmic but weak and infrequent nuclear staining. Biliary excretion of OA was demonstrated, since OA was localized in the lumina of biliary ducts but not within biliary cells.

Administration, Oral↗

PubMed1984

Immunoperoxidase localization of T-2 toxin.

Antibody against T-2 toxin was used for monitoring the fate of T-2 toxin in mice given a single po dose of 11 mg/kg by the peroxidase-antiperoxidase (PAP) method. T-2 toxin was demonstrable in the esophagus from 5 min to about 24 hr postdosing. In the stomach, T-2 toxin was detected within the cytoplasm of intact and injured epithelial cells. In the duodenum, T-2 toxin was primarily localized within the surface epithelium and phagocytic elements (macrophages and neutrophils) of the duodenal lamina propria, especially toward the tips of the villi. Following sloughing of duodenal villous tips, the recovering villous tip epithelial cells frequently showed both cytoplasmic and nuclear T-2 toxin. The jejunum showed weak T-2 toxin within the cytoplasm of villous tip epithelial cells only. The ileum never demonstrated T-2 toxin. Tissue response in the gastrointestinal (GI) tract was characterized by transient edema, marked cytolysis and sloughing, and a subsequent leukocytic invasion of the stomach and proximal small intestine. Evidence of severe gastric and less severe duodenal bleeding was apparent and associated with a marked loss of gastric epithelium and intestinal villous tips. The kidney medulla contained the majority of T-2 toxin stain. T-2 toxin was noted within the distal tubular cells, the cells of the collecting tubules, and the epithelium covering the papilla. T-2 toxin was never demonstrated in any of the hepatic tissue examined.

Administration, Oral↗

PubMed1984

Aflatoxin B1 dihydrodiol antibody: production and specificity.

A specific antibody for 2,3-dihydro-2,3-dihydroxyaflatoxin B1 (AFB1-diol) was prepared, and its reactivity was characterized for the major aflatoxin (AF) B1 (AFB1) metabolites. Reductive alkylation was used to conjugate AFB1-diol to ethylenediamine-modified bovine serum albumin (EDA-BSA) and horseradish peroxidase for use as an immunogen and an enzyme-linked immunosorbent assay (ELISA) marker, respectively. High reactant ratios, 1:5 and 1:10, for AFB1-diol-EDA-BSA (wt/wt) resulted in precipitated conjugates which were poorly immunogenic. However, a soluble conjugate obtained by using a 1:25 ratio of AFB1-diol to EDA-BSA could be used for obtaining high-titer AFB1-diol rabbit antibody within 10 weeks. Competitive ELISAs revealed that the AFB1-diol antibody detected as little as 1 pmol of AFB1-diol per assay. Cross-reactivity of AFB1-diol antibody in the competitive ELISA with AF analogs was as follows: AFB1-diol, 100%; AFB1, 200%; AFM1, 130%; AFB2a, 100%; AFG1, 6%; AFG2, 4%; aflatoxicol, 20%; AFQ1, 2%; AFB1-modified DNA, 32%; and 2,3-dihydro-2-(N7-guanyl)-3-hydroxy AFB1, 0.6%. These data indicated that the cyclopentanone and methoxy moieties of the AF molecule were the primary epitopes for the AFB1-diol antibody. The AFB1-diol competitive ELISA was subject to substantial interference by human, rat, and mouse serum albumins but not by BSA, Tris, human immunoglobulin G, or lysozyme. By using a noncompetitive, indirect ELISA with an AFB1-modified DNA solid phase, a modification level of one AFB1 residue for 200,000 nucleotides could be determined.

Aflatoxin B1↗

PubMed1984

Production and characterization of antibody against aflatoxin Q1.

Antibodies against aflatoxin Q1 (AFQ1) were obtained from rabbits after immunization of either AFQ1-hemisuccinate or AFQ2a conjugated to bovine serum albumin. Both radioimmunoassay and enzyme-linked immunosorbent assaY (ELISA) were used for the determination of antibody titers and specificities. Antibodies obtained from rabbits after immunization with AFQ1-hemisuccinate-bovine serum albumin had the highest affinity to aflatoxin B1, whereas antibodies obtained from rabbits after immunization with AFQ2a-bovine serum albumin bound most effectively with AFQ2a. AFQ2a antibody was selected for the subsequent direct and indirect ELISA for the detection of AFQ1 in biological fluids. When AFQ2a-peroxidase and AFQ2a antibody were used, direct ELISA was able to detect as low as 2 ppb (ng/ml) of AFQ1 spiked in the urine samples that had been subjected to a Sep-Pak cleanup treatment. In indirect ELISA in which the antigen (AFQ2a-bovine serum albumin) was coated to the solid phase followed by reaction with rabbit antibody and goat anti-rabbit immunoglobulin G-peroxidase conjugate, 50-fold less antibody was used without sacrificing sensitivity. Recoveries of AFQ1 added to urine samples (2 to 40 ppb) were 46.3 to 73% and 65.8 to 85.8% for direct and indirect ELISA, respectively.

Aflatoxins↗

PubMed1984

Purification and partial characterization of Flavotoxin A.

A heat-resistant, low-molecular-weight toxin was isolated from semisolid potato dextrose agar medium after inoculation with Flavobacterium farinofermentans sp. nov., which was isolated from fermented corn meal that caused some outbreaks of food poisoning in China. The toxin was purified by solvent partition, Sephadex LH-20 gel filtration, and C-18 reversed-phase column chromatography. Thin-layer chromatography and high-pressure liquid chromatographic methods were developed for the identification and analysis of the toxin. The purified toxin exhibited a single spot in thin-layer chromatography and a single peak in high-pressure liquid chromatography and had adsorption maxima at 232 and 267 nm. Mass spectral analysis indicated a molecular weight of 169 with an experimental formula of C9H13O3. The 50% lethal dose of purified toxin in mice (oral) was less than 6.84 mg/kg, but greater than 0.68 mg/kg. Postmortem examination showed that the mice died of some type of neurological and cardiovascular system toxicity. The name Flavotoxin A is being assigned to the toxin.

Animals↗

PubMed1984

Production and characterization of antibody against diacetoxyscirpenol.

Antibodies against diacetoxyscirpenol (DAS) were obtained from rabbits after immunizing them with hemisuccinate or hemiglutarate derivatives of DAS conjugated to bovine serum albumin (BSA). DAS-hemiglutarate-BSA was found to be a much better immunogen than DAS-hemisuccinate-BSA. Competitive radioimmunoassay revealed that the antisera obtained from rabbits after immunization with DAS-hemiglutarate-BSA showed high specificity toward DAS. The concentrations causing 50% displacement of radioactive DAS by unlabeled DAS, 4-monoacetoxyscirpenol (MAS), and 15-MAS were found to be 1.5, 130, and 300 ng per assay, respectively. Thus, the cross-reactivities for 4-MAS and 15-MAS are ca. 87 and 300 times weaker than that of DAS. Practically no cross-reaction (less than 5% displacement) was observed when deoxynivalenol, T-2 toxin, deoxyverrucarol, and scirpentriol were tested at a concentration of 2,000 ng/ml.

Animals↗

PubMed1984

Production and characterization of antibody against deoxyverrucarol.

Immunization of rabbits with deoxyverrucarol (DOVE) conjugated to bovine serum albumin resulted in antibodies bound with either tritiated DOVE or diacetoxyscirpenol (DAS), but not with T-2 toxin. The affinity of antibodies with DOVE was found to be much higher than with DAS. When [3H] DOVE was used as a marking ligand in the competitive radioimmunoassay, the concentrations causing 50% inhibition of binding radioactivities by unlabeled DOVE, verrucarol, verrucarin A, and 4-monoacetoxyscirpenol were found to be 0.32, 1,070, 9,500, and 10,000 ng per assay, respectively. T-2 toxin, 15-monoacetoxyscirpenol, and deoxynivalenol gave less than 20% inhibition at 10 micrograms per assay. However, when [3H] DAS was used as the marking ligand, the concentrations causing 50% inhibition by DOVE, DAS, and verrucarol were found to be in the 50 to 60 ng per assay range. The antibodies are thus highly specific to DOVE rather than a common trichothecene backbone. The possible use of this antiserum for assay of macrocyclic trichothecenes is discussed.

Animals↗

PubMed1984

Production and characterization of monoclonal antibodies against aflatoxin M1.

By using an indirect enzyme-linked immunosorbent assay, four monoclonal antibodies were selected after fusion of mouse P3-NS1-Ag4-1 myeloma cells with spleen cells isolated from BALB/c mice that had been immunized with aflatoxin M1 (AFM1) conjugated to bovine serum albumin. Two of these antibodies were found to be specific for AFM1 and were designated AMW-1 and AMW-4. The specificities of AMW-1, which had higher affinity to AFM1, were determined by a competitive direct enzyme-linked immunosorbent assay with peroxidase-AFM1 as the marker. The relative cross-reactivity of each toxin (relative to AFM1) with AMW-1, as determined by the amount of aflatoxin necessary to cause 50% inhibition of enzyme activity, was 12, greater than 40, 12, and greater than 40 for B1, B2, G1, and G2, respectively.

Aflatoxin B1↗

PubMed1983

Indirect immunoperoxidase localization of aflatoxin B1 in rat liver.

Aflatoxin B2a (AFB2a) antibody was used as a histochemical probe in the indirect immunoperoxidase localization of aflatoxin B1 (AFB1) bound to rat liver. The efficacy of the indirect method was initially demonstrated by detecting AFB1 covalently bound to DNA in an enzyme-linked immunosorbent assay. AFB1-modified DNA was attached to a polystyrene microtissue culture plate (solid phase) and then subjected to sequential incubation with AFB2a antiserum followed by goat anti-rabbit peroxidase conjugate. Assays for bound peroxidase revealed that the AFB2a antiserum could be diluted 200,000-fold and still yield a signal-to-noise ratio of 10 when compared to an unmodified DNA control. When the same indirect immunoperoxidase protocol was applied to the light-microscopic localization of AFB1 in liver sections of rats treated in vivo with the mycotoxin, bound toxin could be identified in excellent detail in tissues fixed with periodate-lysine-paraformaldehyde and embedded in glycol methacrylate, but was detectable with only poor resolution in unfixed cryostat sections. Peroxidase-positive reactions in hepatocytes typically exhibited strong nuclear and relatively lighter cytoplasmic staining. Greater concentrations of peroxidase-positive hepatocytes were detected in the periportal area than in the area of the central vein, suggesting a circulatory pattern for AFB1 binding in the liver.

Aflatoxin B1↗

PubMed1983

Detection of T-2 toxin by an improved radioimmunoassay.

T-2 toxin in serum, urine, and saline was analyzed by a modified radioimmunoassay procedure. The specimens were added directly to the assay tubes without extraction steps. The reaction between antibody and ligands was optimal at 1 h. Albumin-coated charcoal was used to separate bound from free radioactivity. Quenching, which occurred with hemolyzed specimens, was corrected by a wet oxidation process with 60% perchloric acid and 30% hydrogen peroxide. The shorter incubation times resulted in an assay that takes less than 6 h to complete. The average affinity constant of the antibody (Km) was 1.75 X 10(10) liters/mol. The sensitivity was 1 ng per assay or 10 ng/ml. Among the other trichothecenes tested, only H-T-2 cross-reacted significantly (10.3%).

Antibody Affinity↗

PubMed1983

Toxicity of the Alternaria metabolites alternariol, alternariol methyl ether, altenuene, and tenuazonic acid in the chicken embryo assay.

The effects in the chicken embryo assay of four Alternaria metabolites (alternariol [AOH], alternariol methyl ether [AME], altenuene [ALT], and tenuazonic acid [TA]) were investigated. Administered to 7-day-old chicken embryos by yolk sac injection, AOH, AME, and ALT caused no mortality or teratogenic effect at doses up to 1,000, 500, and 1,000 micrograms per egg, respectively. TA exhibited a calculated 50% lethal dose of 548 micrograms per egg, with no teratogenic effect observed at either lethal or sublethal doses.

Alternaria↗

PubMed1983

Preparation and characterization of acid dehydration products of aflatoxicol.

Treatment of aflatoxicol with acid resulted in the formation of dehydrated derivatives with the 2,3 double bond either hydrated or unaffected. Chromatographic and physicochemical analyses of the purified derivatives revealed that hydration of the 2,3 double bond of aflatoxicol with simultaneous dehydration occurred when sulfuric acid was used. When a weaker acid such as chloroacetic acid was used, no hydration occurred, and 2 isomeric forms of the dehydrated aflatoxicol were obtained. Chicken embryo assay and the Ames test revealed that dehydrated aflatoxicols were much less toxic and mutagenic than aflatoxicol. A mechanism for the dehydration of aflatoxicol is proposed.

Aflatoxins↗

PubMed1982

Ethylenediamine modified bovine serum albumin as protein carrier in the production of antibody against mycotoxins.

The efficiency of conjugation of mycotoxin derivatives containing a carboxylic group or other related functional groups to bovine serum albumin (BSA) by water soluble carbodiimide method, mixed anhydride method and reductive alkylation technique, improved considerably when a modified BSA was used in which the carboxylic groups in the protein were blocked by ethylenediamine (EDA). Due to the increased efficiency, smaller amounts of the haptens were satisfactory for conjugation to EDA-BSA as compared to amounts for the unmodified protein. The mycotoxin-EDA-BSA conjugates were shown to be highly immunogenic in rabbits.

Animals↗

PubMed1982

Kinetics of transformation of aflatoxin B1 into aflatoxin M1 in lactating mouse: an ELISA analysis.

A new enzyme-linked immunosorbent assay (ELISA) was used to study the kinetics of transformation of aflatoxin B1 into aflatoxin M1 in lactating mice. Aflatoxin M1 concentration in the milk samples reached a maximum 30 min after injection of aflatoxin B1 and decreased thereafter. At the maximum time, the levels of aflatoxin M1 in the samples were proportional to the dosages administered. Aflatoxin B1 was also detected in the milk samples but at a lower concentration.

Aflatoxin B1↗