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Biomedical subjects

F S Chu

Publications and source records attributed to F S Chu.

At least 37 records · Page 2Linked to original sources
PubMed1996

Immunochemical methods for fumonisins.

Both monoclonal and polyclonal antibodies have been raised against fumonisins (Fm) and the hydrolyzed toxin in several laboratories. They have been used in several immunoassays and can detect 5 to 5000 ng of FmB1/mL in a "clean" standard solution. However, sample matrices still give severe interferences. Thus, without a cleanup treatment, most immunoassays can only be used as a screening tool for Fm in corn/feed in the 0.5 to 10 ppm range. The antibodies have also been used to generate anti-idiotype antibodies, to use as immunohistological reagents and to make affinity columns that are being used as a cleanup tool. In the present symposium, approaches used to generate antibodies, antibody specificity, sensitivity of various immunoassays and problems encountered in the immunoassays will be reviewed. Approaches used to overcome such problems and methods to improve the sensitivity and specificity of immunoassays of Fm are discussed.

Antibodies↗
PubMed1995

Induction of cytokine mRNAs in mice after oral exposure to the trichothecene vomitoxin (deoxynivalenol): relationship to toxin distribution and protein synthesis inhibition.

The effects of oral exposure to 0, 5, and 25 mg/kg body wt vomitoxin (VT) on cytokine mRNA levels in spleen, Peyer's patches (PP), liver, kidney, and small intestine were evaluated in B6C3F1 mice at 2 and 4 hr postexposure using RT-PCR in conjunction with Southern hybridization analysis. The abundance of mRNAs for several cytokines was increased in VT-exposed mice with maximal effects occurring in the 25 mg/kg group at 2 hr. Specifically, IL-1 beta and IL-6 mRNA levels increased in spleen and PP following exposure to VT. TNF-alpha mRNA levels were markedly elevated in spleen and liver of VT-exposed mice. TGF-beta mRNA was increased in treatment kidneys and to a lesser extent in liver and small intestine. IFN-gamma mRNAs were elevated according to the rank order: spleen > PP > small intestine > liver > kidney, whereas IL-2 mRNAs were increased primarily in spleen and PP. VT had little effect on abundance of mRNAs for the TH2 cytokines, IL-4 and IL-5, or the housekeeping gene, hypoxanthine guanine ribosyl transferase. In order to relate cytokine mRNA abundance to toxin distribution, mice were administered 5 and 25 mg/kg VT body wt containing [3H]VT and tissue levels were monitored over time. Maximum VT molar equivalents for both doses were found at 30 min or 1 hr in all tissues with a rapid clearance following two-compartment kinetics over 24 hr. When effects of oral VT exposure on in vivo protein synthesis at 3 hr postexposure was measured using [14C]leucine uptake, it was found to be inhibited by > or = 20 and > or = 50% in tissues of mice receiving 5 and 25 mg/kg VT, respectively. While recovery was observed in tissues of the 5 mg/kg group at 6 hr, protein synthesis was still significantly inhibited (> or = 70%) at 9 hr for all tissues in the 25 mg/kg group. The results suggest that acute oral VT exposure resulted in the transient elevation of mRNAs for proinflammatory and TH1 cytokines. These effects occurred immediately after peak VT accumulation and concurrently with marked in vivo protein synthesis inhibition.

Administration, Oral↗
PubMed1994

In vitro neutralization of the inhibitory effect of Microcystin-LR to protein phosphatase 2A by antibody against the toxin.

Microcystin-LR (MCYST-LR) was found to be a very potent protein-phosphatase 2A (PP2A) inhibitor at an ID50 as low as 0.1 nM. Comparing to calyculin A and okadaic acid, MCYST-LR was found to be about 100 times stronger than calyculin A and okadaic acid for their inhibition to PP2A. The inhibitory effect on PP2A by MCYST-LR was abolished when antibodies against MCYST-LR were present in the assay system. Polyclonal antibodies were more effective than monoclonal antibodies in reversing the inhibitory effect. A dose-dependent neutralization of the inhibitory effect of MCYST to PP2A by anti-MCYST polyclonal antibodies were observed. Almost 80% of the enzyme activity was restored when as low as 0.012 micrograms of Pab was present. The specific reaction caused by the antibody was evident from an analysis that the antibodies had no effect on reversing the inhibition of PP2A caused by okadaic acid and calyculin A.

Animals↗
PubMed1994

Simultaneous occurrence of fumonisin B1 and other mycotoxins in moldy corn collected from the People's Republic of China in regions with high incidences of esophageal cancer.

A total of 31 corn samples collected from households in the counties of Cixian and Linxian of the People's Republic of China, where high incidences of esophageal cancer have been reported, were analyzed for fumonisin B1 (FB1), aflatoxin, and total trichothecene mycotoxins. High levels of FB1 (18 to 155 ppm; mean, 74 ppm) were found in 16 of the samples that showed heavy mold contamination. FB1, at lower levels (20 to 60 ppm; mean, 35.3 ppm), was also found in 15 samples, collected from the same households, that did not show any visible mold contamination. The levels of aflatoxin in the samples were low (1 to 38.4 ppb; mean, 8.61 ppb). High levels of total type-A trichothecenes were also found in the moldy corn samples (139 to 2,030 ppb; mean, 627 ppb). Immunochromatography of selected samples revealed that these samples contained T-2 toxin, HT-2 toxin, iso-neosolaniol, monoacetoxyscirpenol, and several other type-A trichothecenes. The concentration of total type-B trichothecenes in 15 moldy corn samples was in the range of 470 to 5,826 ppb (mean, 2,359 ppb). High levels (3.7 to 5.0 mg/g) of FB1 were produced in corn in the laboratory by five Fusarium moniliforme strains isolated from the moldy corn. These fungi were also capable of forming various nitrosamines (5 to 16 micrograms per flask) in the presence of nitrate and precursor amines.(ABSTRACT TRUNCATED AT 250 WORDS)

China↗
PubMed1993

Cloning and characterization of a cDNA from Aspergillus parasiticus encoding an O-methyltransferase involved in aflatoxin biosynthesis.

Aflatoxins are polyketide-derived secondary metabolites produced by the fungi Aspergillus flavus and Aspergillus parasiticus. Among the catalytic steps in the aflatoxin biosynthetic pathway, the conversion of sterigmatocystin to O-methylsterigmatocystin and the conversion of dihydrosterigmatocystin to dihydro-O-methylsterigmatocystin are catalyzed by an S-adenosylmethionine-dependent O-methyltransferase. A cDNA library was constructed by using RNA isolated from a 24-h-old culture of wild-type A. parasiticus SRRC 143 and was screened by using polyclonal antiserum raised against a purified 40-kDa O-methyltransferase protein. A clone that harbored a full-length cDNA insert (1,460 bp) containing the 1,254-bp coding region of the gene omt-1 was identified by the antiserum and isolated. The complete cDNA sequence was determined, and the corresponding 418-amino-acid sequence of the native enzyme with a molecular weight of 46,000 was deduced. This 46-kDa native enzyme has a leader sequence of 41 amino acids, and the mature form of the enzyme apparently consists of 377 amino acids and has a molecular weight of 42,000. Direct sequencing of the purified mature enzyme from A. parasiticus SRRC 163 showed that 19 of 22 amino acid residues were identical to the amino acid residues in an internal region of the deduced amino acid sequence of the mature protein. The 1,460-bp omt-1 cDNA was cloned into an Escherichia coli expression system; a Western blot (immunoblot) analysis of crude extracts from this expression system revealed a 51-kDa fusion protein (fused with a 5-kDa beta-galactosidase N-terminal fragment).(ABSTRACT TRUNCATED AT 250 WORDS)

Aflatoxins↗
PubMed1993

Rectal perforation after barium enema in a patient with cytomegalovirus colitis--a case report.

The very rare complication of rectal perforation during the barium enema of an immunocompetent Chinese man with cytomegalovirus (CMV) colitis is described. Rectal biopsy performed over three weeks earlier was unlikely to have been related to the perforation. Histopathological examination of the colonic mucosa showed features of active chronic colitis and CMV infection. After treatment with Ganciclovir, there was cessation of symptoms with eradication of the virus on repeat biopsy samples. It is suggested that presence of CMV colitis predisposed to rectal perforation. The clinical presentation, course of disease and response to treatment supported the concept of CMV as a primary aetiological agent. This complication should be borne in mind when performing barium enemas in patients with clinical features of colitis, especially if they are immunocompromised.

Aged↗
PubMed1992

Variation in regulation of aflatoxin biosynthesis among isolates of Aspergillus flavus.

Two new phenotypes of Aspergillus flavus which exhibit novel patterns of aflatoxin production have been identified and characterized. In one of the new variants of A. flavus, aflatoxin is made in the absence of carbohydrate and concomitantly with growth, without a lag period. A second variant did not produce aflatoxin in the presence or absence of carbohydrate. Chemical mutagenesis of this nonaflatoxigenic strain resulted in mutant strains which produced aflatoxin on carbohydrate-containing media. The aflatoxin production pattern observed in these mutants resembled the typical production scheme, with a lag period through log phase growth.

Aflatoxins↗
PubMed1992

Molecular cloning of genes related to aflatoxin biosynthesis by differential screening.

A differential hybridization strategy was used to clone genes associated with aflatoxin biosynthesis. A genomic library, formed between nuclear DNA and the pUC19 plasmid, was screened with three different cDNA probes by the colony hybridization procedure. Nineteen clones were selected; all were positively correlated with and presumably enriched with genes associated with aflatoxin production. Some of these clones were further characterized by using them as probes in Northern (RNA blot) hybridizations. Five clones hybridized strongly with some polyadenylated RNAs formed during the transition to or during idiophase when aflatoxin was produced. However, little or no corresponding hybridization occurred with polyadenylated RNAs formed in early and mid-log growth phase. Two of the clones were further used as probes to hybridize with polyadenylated RNAs formed under aflatoxin-permissive and nonpermissive temperatures. Hybridization occurred with RNA species formed under the permissive temperature only.

Aflatoxins↗
PubMed1992

Recent progress on analytical techniques for mycotoxins in feedstuffs.

Analysis of mycotoxins in feedstuffs is a difficult task because only trace amounts of the toxins are present in the sample. However, rapid progress in the area of mycotoxin analysis has been made during the last few years. Simplified sample cleanup protocols and new chromatographic methods, especially HPLC, have been developed. New, more sensitive and versatile instruments such as high-resolution mass spectrometry (MS) and gas chromatography/tandem MS/MS are coming to the market. After 15 yr of laboratory research, immunoassay techniques have gained more acceptance as analytical tools for mycotoxins. Several immunoassay kits for mycotoxins are currently available. The development of these new techniques and their application for monitoring various mycotoxins in foods and feeds are described in this review.

Animal Feed↗
PubMed1991

Production and characterization of antibodies against nivalenol tetraacetate.

Antibody against nivalenol tetraacetate (tetra-Ac-NIV) was prepared by immunization of rabbits with triacetyl-15-pimelate-NIV conjugated to bovine serum albumin. By using tritiated tetra-Ac-NIV as the test ligand, antibody titers were demonstrated as early as 4 weeks after immunization. Useful antibody for radioimmunoassay (RIA) of tetra-Ac-NIV was obtained 7 weeks after immunization, with one booster injection. Results of competitive RIA revealed that the antibody was most specific to tetra-Ac-NIV. The relative cross-reactivity of this antibody with tetra-Ac-NIV, deoxynivalenol triacetate, and neosolaniol triacetate was found to be 100, 2.2, and less than 1, respectively. Practically no cross-reaction was found with deoxynivalenol, fusarenon X, and NIV. The detection limit for tetra-Ac-NIV by RIA was about 5.0 ng/ml (0.5 ng per assay). The use of this antibody for quantitation of NIV in cereals after acetylation of sample extracts is proposed.

Acetylation↗
PubMed1989

Production and characterization of antibodies cross-reactive with major aflatoxins.

Antibodies cross-reactive with 4 major aflatoxins were demonstrated three weeks after immunization of rabbits with an immunogen which was prepared by conjugating aflatoxin B3 to bovine serum albumin. Aflatoxin B3 was first converted to its hemisuccinate before conjugation to the protein. Tritiated aflatoxin B1 (AFB1) was used as the marker ligand both for antibody titer determination as well as for analysis of antibody specificity. Competitive RIA revealed that the antibodies have good cross-reactivity with aflatoxins B1, B2, G1, and G2 when tritiated AFB1 was used as the marker ligand. The concentrations causing 50% inhibition of binding of 3H-AFB1 to the antibodies by unlabeled aflatoxins B1, B2, G1, G2 and B3 were found to be 0.25, 3.34, 0.32, 4.0 and 0.53 ng/assay, respectively. The antibodies could be used for simultaneous analysis of aflatoxins B1 and G1, two of the most important toxic metabolites produced by Aspergillus flavus and A. parasiticus.

Aflatoxin B1↗
PubMed1989

Production and characterization of antibodies against microcystins.

Antibodies against a microcystin (MCYST) leucine-arginine variant (MCYST-LR) were demonstrated 4 weeks after immunization of rabbits with either MCYST-LR-polylysine- or MCYST-LR-ethylenediamine-modified bovine serum albumin. A radioimmunoassay (RIA), a direct competitive enzyme-linked immunosorbent assay (ELISA), and an indirect competitive ELISA were developed for characterization of the antibodies. Indirect ELISA and RIA revealed that MCYST-LR-ethylenediamine-bovine serum albumin was a better immunogen. Competitive RIA and direct ELISA revealed that the antibodies had good cross-reactivities with an MCYST-arginine-arginine variant (MCYST-RR), MCYST-LR, an MCYST-tyrosine-arginine variant (MCYST-YR), and nodularin (NODLN); but they had lower reactivities with variants MCYST-leucine-tyrosine (MCYST-LY) and MCYST-leucine-alanine (MCYST-LA). The antibodies did not cross-react with ozonolyzed MCYST-LR. The concentrations causing 50% inhibition of binding of reduced MCYST-LR to the antibodies by MCYST-RR, MCYST-LR, MCYST-YR, NODLN, MCYST-LA, and MCYST-LY in the RIA were 43, 105, 112, 503, 671, and 1,920 ng/ml, respectively. The concentrations causing 50% inhibition of binding of MCYST-LR-horseradish peroxidase to the antibodies by MCYST-RR, MCYST-LR, MCYST-YR, NODLN, MCYST-LY, and MCYST-LA in the ELISA were 1.75, 2.2, 3.4, 4.6, 50, and 114 ng/ml, respectively.

Animals↗
PubMed1988

Uptake and metabolism of T-2 toxin in relation to its cytotoxicity in lymphoid cells.

The sensitivity of lymphoid cells to the cytotoxic effects of T-2 toxin (T-2) varies according to their degree of differentiation. To understand the mechanisms of these variations, the uptake and the metabolism of T-2 in susceptible (human lymphoma Daudi and phytohaemagglutinin-stimulated murine lymphocytes) and resistant (human leukaemia KE37 and REH) cells were studied in culture. When cells were incubated with [3H]T-2 a significant increase in the quantity of T-2 associated with the cell occurred during the first 30 min, this increased further from 10-16 hr, and decreased after 24 hr. Daudi and REH cells took up 20 and 3% of the T-2 present in the medium, respectively. Metabolites, extracted from the culture medium and from cells, were analysed by the thin-layer chromatography. The products were identified by comparison with standards for T-2 tetraol, T-2 triol, HT-2 toxin, neosolaniol and T-2. Qualitatively, similar metabolic pathways were found in all cells examined. The presence of these metabolites demonstrated that T-2 was taken up by these cells. A correlation existed between the relative sensitivities of the cells toward T-2 and the amount of intracellular T-2 and/or metabolites. It is thought that differences in the kinetics of uptake and processing of T-2 account for the known differences in cellular sensitivities to the toxin.

Cells, Cultured↗
PubMed1988

Production and characterization of a monoclonal antibody cross-reactive with most group A trichothecenes.

A monoclonal antibody cross-reactive with most group A trichothecenes was produced by fusion of P3/NS-1/1-AG4-1 myeloma cells with spleen cells isolated from a BALB/c mouse that had been immunized with 3-acetyl-neosolaniol-hemisuccinate conjugated to bovine serum albumin. One stable clone, H159B1D5, which produced monoclonal antibody that bound with both T-2 toxin and diacetoxyscirpenol (DAS) was obtained after subcloning. Enzyme-linked immunosorbent assay (ELISA) revealed that the antibody belongs to the immunoglobulin G1 (kappa chain) isotype and had binding constants of 2.81 x 10(9), 1.05 x 10(9), and 1.57 x 10(8) liters per mole for T-2 tetraol tetraacetate, T-2 toxin, and DAS, respectively. The relative cross-reactivities of the antibody with T-2 tetraol tetraacetate, T-2 toxin, and DAS were 200, 100, and 20, respectively, with tritiated T-2 toxin as the marker ligand. The relative cross-reactivities for the above toxins were 667, 100, and 73, respectively, with tritiated DAS as the marker ligand. No cross-reaction with HT-2 and deoxynivalenol triacetate was observed in either system. By using this monoclonal antibody, an indirect ELISA for analysis of T-2 toxin was also developed. The linear portion of the standard curve for analysis of T-2 toxin in each analysis by radioimmunoassay and ELISA was in the range of 0.1 to 2 ng and 0.05 to 1.0 ng, respectively.

Animals↗
PubMed1987

Correlation of dietary aflatoxin B1 levels with excretion of aflatoxin M1 in human urine.

Corn and peanut oil (total, 253 samples) were collected from 32 households in Fushui county of the Guangxi autonomous region of the People's Republic of China, where high liver cancer incidence has been reported, every day over a period of 1 week and analyzed for aflatoxin B1 (AFB). A total of 252 urine samples were collected simultaneously from the residents in the households which were shown to have consumed AFB and were analyzed for aflatoxin M1 (AFM) by a competitive direct enzyme-linked immunosorbent assay. A good correlation between total dietary AFB intake and total AFM excretion in human urine was observed during a 3-day study. A regression equation of 0.143 plus 0.0135 multiplied by the amount of AFB consumed was observed. Between 1.23 and 2.18% of dietary AFB was found to be present as AFM in human urine. A good correlation was also observed between the AFB concentration in corn and the AFM concentration in human urine. The results suggest that analysis of AFM in urine by enzyme-linked immunosorbent assay could be used as an index for human exposure of AFB in an extensive epidemiological study.

Aflatoxin B1↗