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At least 19 recordsLinked to original sources
PubMed1999

Combined secondary enrichment of primary enrichment broths increases Listeria detection.

The efficacy of combining dual primary enrichment cultures into a single secondary broth was evaluated for detecting Listeria in naturally contaminated meats and environmental samples obtained from dairy processing plants. A total of 336 samples were tested using University of Vermont modified Listeria enrichment broth (UVM) and Listeria repair broth containing selective agents (LRBS) as primary enrichment media. Eighty samples (23.8%) yielded Listeria by at least one method. Neither primary enrichment broth was significantly better (P>0.05) than the other in identifying Listeria-positive samples. UVM media, when used as a primary enrichment broth, identified 66 Listeria-positive samples, while the use of LRBS as a primary enrichment broth identified 65 Listeria-positive samples. Listeria detection improved significantly (P<0.01) when two primary enrichment media were used for sample analysis. It is not clear whether this improvement was due to simply replicating the primary enrichment or to the particular pair of primary enrichment media used. The use of a dual secondary enrichment procedure was better (P<0.05) than the use of either individual primary enrichment medium alone. The overall rate of recovery increased from 81.3 to 82.5% for single secondary enrichment to 93.8% using a dual secondary enrichment technique. Analysis of results obtained when combining two independent isolation methods versus combining two primary enrichment media into one single secondary enrichment broth indicated that there was no significant difference (P>0.05) in either procedure. Inoculum size (0.1 ml versus 0.2 ml) did not have an effect on the overall rate of recovery. The procedure developed increased the sensitivity of testing while decreasing the potential workload associated with an increase in enrichment procedures.

Animals↗
PubMed1997

Construction and characterization of a full length-enriched and a 5'-end-enriched cDNA library.

Using 'oligo-capped' mRNA [Maruyama, K., Sugano, S., 1994. Oligo-capping: a simple method to replace the cap structure of eukaryotic mRNAs with oligoribonucleotides. Gene 138, 171-174], whose cap structure was replaced by a synthetic oligonucleotide, we constructed two types of cDNA library. One is a 'full length-enriched cDNA library' which has a high content of full-length cDNA clones and the other is a '5'-end-enriched cDNA library', which has a high content of cDNA clones with their mRNA start sites. The 5'-end-enriched library was constructed especially for isolating the mRNA start sites of long mRNAs. In order to characterize these libraries, we performed one-pass sequencing of randomly selected cDNA clones from both libraries (84 clones for the full length-enriched cDNA library and 159 clones for the 5'-end-enriched cDNA library). The cDNA clones of the polypeptide chain elongation factor 1 alpha were most frequently (nine clones) isolated, and more than 80% of them (eight clones) contained the mRNA start site of the gene. Furthermore, about 80% of the cDNA clones of both libraries whose sequence matched with known genes had the known 5' ends or sequences upstream of the known 5' ends (28 out of 35 for the full length-enriched library and 51 out of 62 for the 5'-end-enriched library). The longest full-length clone of the full length-enriched cDNA library was about 3300 bp (among 28 clones). In contrast, seven clones (out of the 51 clones with the mRNA start sites) from the 5'-end-enriched cDNA library came from mRNAs whose length is more than 3500 bp. These cDNA libraries may be useful for generating 5' ESTs with the information of the mRNA start sites that are now scarce in the EST database.

Animals↗
PubMed1996

Protein tyrosine kinase inhibitors suppress the production of nitric oxide in mixed glia, microglia-enriched or astrocyte-enriched cultures.

Nitric oxide (NO) produced by glial cells has been implicated in the neuropathogenesis of various diseases. However, the signaling transduction pathway(s) for the production of NO in these cells is not well understood. To test whether protein tyrosine kinases (PTKs) are required for signaling events of NO production in glial cells, this study examined the effects of genistein and tyrphostin A25, two potent inhibitors of PTKs, on the production of NO in mouse primary mixed glia, microglia-enriched or astrocyte-enriched cultures exposed to lipopolysaccharide (LPS) or a combination of LPS and interferon-gamma (IFN gamma). LPS induced a dose-dependent increase in NO production from the mixed glia cultures. The LPS-induced NO production was significantly enhanced by stimulating the cells with IFN gamma. Genistein or tyrphostin A25 inhibited the production of NO in both LPS- and IFN gamma/LPS-stimulated mixed glia cultures. The production of NO in the stimulated microglia-enriched or astrocyte-enriched cultures was also inhibited by tyrphostin A25. To verify the cellular sources of NO, immunocytochemical staining of inducible NO synthase (iNOS) was followed by staining with the microglia marker Mac-1 or the astrocyte marker glial fibrillary acid protein (GFAP) in microglia-enriched or astrocyte-enriched cultures. The expression of iNOS and the production of NO in microglia-enriched cultures were significantly higher than those in the identically stimulated astrocyte-enriched cultures. These results demonstrate that PTKs are involved in the signaling events of LPS-induced NO production in microglia and astrocytes, and that microglia are more responsive than astrocytes to stimuli which induce NO. These results may provide insights into therapeutic interventions in the pathway for NO production in the brain.

Analysis of Variance↗
PubMed1987

Comparison of alcohol shock enrichment and selective enrichment for the isolation of Clostridium difficile.

Two enrichment methods were compared for their ability to recover Clostridium difficile from stool samples. One method used selective enrichment in an antibiotic-containing broth followed by detection with a latex particle agglutination (LPA) reagent. The other used enrichment in a non-selective broth following treatment of the specimen with alcohol. With clinical specimens enrichment culture was significantly more successful at detecting C. difficile than direct plating. Alcohol shock enrichment was twice as effective as direct culture, while selective broth enrichment was three times more effective. The use of LPA for screening selective enrichment broths for C. difficile should prove a cost-effective measure as only positive broths (about 20%) require subculture for confirmation.

Bacteriological Techniques↗
PubMed2005

How do giant pandas (Ailuropoda melanoleuca) respond to varying properties of enrichments? A comparison of behavioral profiles among five enrichment items.

This study investigated the responses of 14 giant pandas (Ailuropoda melanoleuca) to 5 enrichment items (3 manipulable objects, 2 feeding devices) that possess different properties affording varying behavioral opportunities. Enrichment items were quantitatively described by using 12 properties. Pandas displayed no preferences among the 5 enrichment treatments, all of which had similar positive effects on behavioral measures of well-being (behavioral diversity, stereotypy). However, each item promoted a distinct behavioral profile, readily mapped onto the enrichment's properties. These results suggest that it may be important to stimulate behavioral diversity but that specific behaviors or enrichment properties are not required. The unique behavioral topography seen with these enrichments also suggests that providing multiple enrichments with varying properties will maximize overall behavioral diversity.

Animals↗
PubMed1984

Tropomyosin-enriched and alpha-actinin-enriched microfilaments isolated from chicken embryo fibroblasts by monoclonal antibodies.

Antitropomyosin and anti-alpha-actinin monoclonal antibodies have been used to isolate two classes of microfilaments, i.e., tropomyosin-enriched and alpha-actinin-enriched microfilaments, respectively, from cultured chicken embryo fibroblasts. Electron microscopic studies of the isolated tropomyosin-enriched microfilaments showed periodic localization of tropomyosin along the microfilaments, with a 35-nm repeat. On the contrary, the isolated alpha-actinin-enriched microfilaments showed no obvious periodicity. Many individual alpha-actinin-enriched microfilaments with length greater than 1 micron (ranging from 1 to 10 microns) were aggregated by anti-alpha-actinin monoclonal antibodies. Both of the isolated microfilaments had the ability to activate the Mg2+-ATPase activity of skeletal muscle myosin, although different extents of activation were observed. These two classes of microfilaments also differed in their protein composition. Molar ratios of major identifiable proteins in the isolated microfilaments were alpha-actinin(dimer):actin(monomer):tropomyosin(dimer) = less than 0.02:8.06:1.00 for tropomyosin-enriched microfilaments and 0.44:13.91:1.00 for alpha-actinin-enriched microfilaments. By two-dimensional gel analysis of the isolated microfilaments, we have found seven spots which possess typical tropomyosin properties including pI 4.5, immunological cross-reaction, lack of proline and tryptophan, and heat stability. Pulse-chase experiments suggested that the assembly of microfilament-associated proteins, at least for alpha-actinin and tropomyosins, was coordinately regulated by the assembly of actin into microfilaments.

Actinin↗
PubMed1994

Effects of linoleate-enriched and oleate-enriched diets in combination with alpha-tocopherol on the susceptibility of LDL and LDL subfractions to oxidative modification in humans.

This report describes the effects of feeding linoleate- or oleate-enriched diets to subjects who were concurrently taking 1200 mg/d of alpha-tocopherol on the susceptibility of low-density lipoprotein (LDL) and buoyant and dense LDL subfractions to oxidation. LDL isolated from subjects who consumed linoleate-enriched diets was more susceptible to copper-mediated oxidation, as measured by formation of conjugated dienes and lipid peroxides and loss of unsaturated fatty acids, compared with LDL isolated from subjects who consumed their usual or oleate-enriched diets. In all subjects, buoyant LDL had a higher content of alpha-tocopherol per particle and a lower 18:2 to 18:1 ratio and was considerably more resistant to oxidation than dense LDL. Although dense LDL from all groups had comparable alpha-tocopherol levels, dense LDL from the linoleate group was most susceptible to oxidation, followed by that from the standard diet, whereas dense LDL isolated from the oleate diet group was most resistant. In summary, high dosages of alpha-tocopherol did not prevent enhanced susceptibility to oxidation of LDL isolated from subjects fed linoleate-enriched diets. Furthermore, dense LDL was more susceptible to oxidation than was buoyant LDL, and this effect was greatly exaggerated in the dense LDL isolated from subjects fed linoleate-enriched diets. Conversely, dense LDL isolated from subjects fed oleate-enriched diets was the most protected. If oxidation of LDL is important in the pathogenesis of atherosclerosis, then these data suggest that in people with increased amounts of small, dense LDL, dietary enrichment in oleic acid may decrease the susceptibility of their LDL to oxidation.

Adult↗
PubMedDate not supplied

Salmonella in foods: new enrichment procedure for TECRA Salmonella Visual Immunoassay using a single rv(R10) only, TT only, or dual rv(R10) and TT selective enrichment broths (AOAC official method 998.09): collaborative study.

A collaborative study was conducted to compare a new enrichment procedure for the TECRA Salmonella Visual Immunoassay (TSVIA) with the reference method given in the U.S. Food and Drug Administration's Bacteriological Analytical Manual (7th Ed.). Three food types (milk powder, pepper, and soy flour) were analyzed in Australia and 3 food types (milk chocolate, dried egg, and raw turkey) were analyzed in the United States. Thirty-eight collaborators participated in the study. The TECRA method was evaluated using both Rappaport-Vassiliadis R10 (RV(R10)) and tetrathionate (TT) broths for selective enrichment. M broth cultures arising from each of the 2 selective enrichment broths were tested in the TSVIA using 2 individual wells, one for each selective broth, and a single well to test the pooled selective enrichment broths. The results for the pooled enrichment broths were reported elsewhere. This study presents the results for the use of single enrichment broths, i.e., RV(R10) only or TT only, with the TSVIA. No significant differences (p > 0.05) were observed for the pairwise comparison of the proportion of positive samples for either RV(R10) or TT used as a single enrichment broth for the TSVIA with that for the reference method.

Animals↗
PubMedDate not supplied

Detection of Salmonella in dry foods using refrigerated pre-enrichment and enrichment broth cultures: interlaboratory study.

An interlaboratory study was performed in 11 laboratories to validate the use of pre-enrichment and tetrathionate brilliant green (TBG35) and selenite cystine (SC35) enrichment cultures refrigerated 72 h at 2-5 degrees C for greater analytical flexibility in the detection of Salmonella in dry foods. Productivities of refrigerated pre-enrichment and enrichment cultures were compared with that of the AOAC/Bacteriological Analytical Manual (BAM) procedure using 4 food types: whole egg powder, milk chocolate, animal feed, and instantized skim milk powder. Uninoculated and inoculated samples were included in each food group. There was complete agreement between the results obtained by the standard AOAC/BAM procedure and the 2 refrigeration procedures. Of 660 samples tested, the AOAC/BAM procedure identified 393 contaminated samples that were readily detected from the corresponding refrigerated pre-enrichment cultures and from the combined productivity of homologous refrigerated TBG35 and SC35 cultures. Refrigeration (72 h) of pre-enrichment or enrichment cultures for greater analytical flexibility and laboratory productivity in the examination of dry foods is under review for adoption by AOAC International.

Animal Feed↗
PubMed2001

Immunoseparation of sphingolipid-enriched membrane domains enriched in Src family protein tyrosine kinases and in the neuronal adhesion molecule TAG-1 by anti-GD3 ganglioside monoclonal antibody.

Rat cerebellar granule cells differentiated in culture were fed [1-(3)H]sphingosine, allowing the metabolic radiolabelling of all cell sphingolipids and phosphatidylethanolamine. A detergent-insoluble sphingolipid-enriched membrane fraction, containing about 60% of cell sphingolipids, but only trace amounts of phosphatidylethanolamine, was prepared from [1-(3)H]sphingosine-fed cells by sucrose gradient centrifugation. This fraction was enriched in the Src family protein tyrosine kinases c-Src, Lyn and Fyn and in the GPI-anchored neuronal adhesion molecule TAG-1. The cell lysate and the sphingolipid-enriched membrane fraction were subjected to immunoprecipitation with anti-GD3 ganglioside monoclonal antibody R24, under experimental conditions designed to preserve the integrity of the domain. The radioactive lipid composition of the immunoprecipitates obtained from the cell lysate and from the sphingolipid-enriched fraction were very similar, and closely resembled the sphingolipid composition of the whole sphingolipid-enriched membrane fraction. In fact, the immunoprecipitates contained, together with GD3 ganglioside, all cell glycosphingolipids and sphingomyelin, whereas they did not contain phosphatidylethanolamine. Moreover, cholesterol and phosphatidylcholine were detected in the immunoprecipitates by qualitative TLC analysis followed by colourimetric visualization. c-Src, Lyn, Fyn and TAG-1 were associated with the anti-GD3 antibody immunoprecipitate. These proteins were not detected in the immunoprecipitates obtained under experimental conditions different from those designed to preserve the integrity of the domain. These data suggest that a membrane domain containing cholesterol, phosphatidylcholine, sphingolipids and proteins can be separated from the total cell membranes by anti-GD3 antibody immunoprecipitation, and that the association of c-Src, Fyn, Lyn, and TAG-1 with the sphingolipid-enriched domain is mediated by the interaction with a complex lipid environment, rather than by specific interactions with a single sphingolipid species.

Animals↗
PubMed2025

Gene behaviors-based network enrichment analysis and its application to reveal immune disease pathways enriched with COVID-19 severity-specific gene networks.

MOTIVATION: Gene network analysis is essential for understanding the complex mechanisms underlying diseases, which often involve disruptions in molecular networks rather than individual genes. Despite the availability of large-scale omics datasets and computational tools for gene network analysis, interpretation of the biological relevance of these extensive networks remains challenging. RESULTS: We propose a novel computational strategy, gene behaviors-based network enrichment analysis, which systematically identifies functional pathways enriched in phenotype-specific gene networks. Our novel method incorporates comprehensive network characteristics, i.e. gene expression levels, edge strengths, and structural patterns of edges, to rank genes based on activity and assess pathway enrichment, effectively identifying functional pathways enriched within these networks. Through simulation studies, our strategy demonstrated superior performance compared with that of existing methods in identifying enriched pathways. We applied this strategy to whole-blood RNA-seq data from 1102 COVID-19 samples provided by the Japan COVID-19 Task Force. The analysis revealed immune disease pathways enriched with COVID-19 severity-specific gene networks, including "Systemic lupus erythematosus" in asymptomatic and severe samples and "Inflammatory bowel disease," "Primary immunodeficiency," and "Rheumatoid arthritis" in mild samples. Key biomarkers of COVID-19, such as CXCL8, S100A9, and HLA class I genes, have been identified as critical hub genes and the main players within these networks. AVAILABILITY AND IMPLEMENTATION: Code is available in Figshare (https://doi.org/10.6084/m9.figshare.29093648.v3).

COVID-19↗
PubMed2004

To enrich or not to enrich: providing shelter does not complicate handling of laboratory mice.

Environmental enrichment (EE) is used in laboratory animal housing to provide stimuli exceeding those of barren cages and is intended to improve the welfare of captive animals. It is argued that when laboratory mice can routinely retreat in sheltering objects when humans are present, they do not habituate to humans and continue to shy away, thereby increasing the time needed for husbandry and testing procedures. To this date very limited research has been carried out to determine whether providing EE in the form of shelter interferes with the habituation of mice to humans and thus complicates catching and handling them. We housed 20 FVB (inbred) and 20 NMRI (outbred) male mice in standard cages and another 20 FVB and 20 NMRI male mice in cages enriched with two PVC conduits. When the mice were 10 weeks old, measurements of food and water consumption, weight, latency of catching, and a behavior score in response to handling during a sham subcutaneous injection were performed weekly for 4 consecutive weeks. Food and water consumption and weight were influenced by strain, but the presence of EE in the home cage did not affect these parameters as much. Outbred mice ate, drank, and weighed more than did the inbred animals, but they did not significantly gain weight during the course of the 4 testing weeks. Cage enrichment in the form of PVC conduits decreased the time needed to catch outbred animals and did not increase the time needed to catch mice from the inbred strain. Furthermore, no differences in resistance to being held during the sham injection could be detected between animals from the enriched versus non-enriched group. These results indicate that EE in the form of sheltering objects does not complicate catching or handling mice and that allowing access to enrichment in the laboratory cage, which has been shown to have positive effects on welfare, does not interfere with the management or cost of laboratory animals.

Animal Husbandry↗
PubMed1993

A simple technique for the rapid enrichment of class and subclass hybridoma switch variants. A 1000-fold enrichment in half the time, for half the cost.

Switching parental hybrids in vitro to downstream switch variant clones producing more desirable monoclonal antibodies (MoAbs) requires either labor intensive and time consuming subcloning techniques, or fluorescence activated sorting of the desired clones. We tested the hypothesis that enrichment of downstream switch variant clones might be achieved by selective lysis of upstream hybridoma cells followed by expansion of the enriched downstream clone. Using a parental hybridoma with surface and secretory IgM, we attempted to enrich downstream switch variant clones producing class (IgG) and subclass (IgG1 or IgG2a) MoAbs. Enrichment of downstream IgG, IgG1 and IgG2a MoAb-producing switch variants was achieved by single or repeated antibody-dependent, complement-mediated lysis of the upstream IgM-bearing parental hybridoma cells followed by limited subcloning. Two exposures of parental hybridoma cells to lysis followed by plating at 100 cells/well enriched the frequency of switch variants up to 1235-fold, enabling the development of IgG1 or IgG2a-producing subclones exhibiting high yield antibody production. Using this protocol, production time and costs were reduced by > 50% when compared to the standard technique. This novel technique for the rapid isolation and expansion of switch variant clones should be ideal for most laboratories, particularly those without access to cell sorting capabilities.

Animals↗
PubMed1970

Comparative studies on mitochondria isolated from neuron-enriched and glia-enriched fractions of rabbit and beef brain.

Fractions enriched in neuronal and glial cells were obtained from dispersions of whole beef brain and rabbit cerebral cortex by large-scale density gradient centrifugation procedures. The fractions were characterized by appropriate microscopic observation. Mitochondria were then isolated from these fractions by differential centrifugation of their homogenates. The two different types of mitochondria were characterized with respect to certain enzyme activities, respiratory rate, rate of protein synthesis, and their buoyant density in sucrose gradients. The mitochondria from the neuron-enriched fraction were distinguished by a higher rate of incorporation of amino acids into protein, higher cytochrome oxidase activity, and a higher buoyant density in sucrose density gradients. Mitochondria from the glia-enriched fraction showed relatively high monoamine oxidase and Na(+)- and K(+)-stimulated ATPase activities. The rates of oxidation of various substrates and the acceptor control ratios did not differ appreciably between the two types of mitochondria. The difference in the buoyant density of mitochondria isolated from the neuron-enriched and glia-enriched cell fractions was utilized in attempts to separate neuronal and glial mitochondria from the mixed mitochondria obtained from whole brain homogenates in shallow sucrose gradients. The appearance of two peaks of cytochrome oxidase, monoamine oxidase, and protein concentration in such gradients shows the potential feasibility of such an approach.

Animals↗
PubMed1998

Enrichment of hen eggs with n-3 long-chain fatty acids and evaluation of enriched eggs in humans.

Eggs enriched with n-3 polyunsaturated fatty acids (PUFAs) were produced by hens fed diets containing fish oil or a combination of fish and vegetable oils. In a sensory evaluation, 78 untrained volunteers could not distinguish between ordinary and enriched eggs. Storage life was also not significantly different between egg types. A food intake survey of 4 groups of 14 subjects each who consumed 7 eggs/wk for 24 wk showed that intakes of the major dietary components were not significantly different for 4 different egg types. Mean (n = 56) plasma cholesterol and triacylglycerol concentrations were not significantly different at the start and finish of the study. Body weight and HDL concentrations increased during the study (P < 0.05). For the last 2 wk of the experiment (weeks 23-24), mean egg consumption was increased from a total of 14 to a total of 21 eggs, resulting in a small increase in plasma triacylglycerols only. There were no significant differences (P > 0.05) in body weight, blood pressure, or plasma lipid components among treatment groups consuming the 4 different egg types. Blood samples taken after 16 and 22 wk from fasted subjects showed significant increases in eicosapentaenoic acid, docosahexaenoic acid, and total n-3 PUFAs in subjects consuming enriched eggs compared with controls. In addition, the ratio of n-6 to n-3 PUFAs in plasma was significantly reduced from 12.2:1 to 6.5-7.7:1 in subjects consuming enriched eggs compared with controls. Consumption of only one enriched egg daily can contribute substantially to the recommended daily intake of n-3 PUFAs.

Adult↗
PubMed1999

Sphingomyelins of rat liver: biliary enrichment with molecular species containing 16:0 fatty acids as compared to canalicular-enriched plasma membranes.

We harvested canalicular-enriched plasma membranes of hepatocytes and collected fistula bile from male rats and isolated the sphingomyelins. Following sphingomyelinase hydrolysis, we identified the sphingomyelin molecular species on the basis of their benzoylated ceramide derivatives employing high performance liquid chromatography. Sphingomyelin constitutes </=3% of total biliary phospholipids (which are mostly sn-1 16:0 long-chain phosphatidylcholines) and approximately 30% of canalicular-enriched membranes. In both cases, the principal molecular species were composed of 16:0, 18:0, 20:0, 22:0, 23:0, 24:0, 24:1 and 24:2 fatty acid classes. However, the 16:0 fatty acid species was enriched in biliary sphingomyelin to a significantly greater degree than in sphingomyelins of canalicular-enriched plasma membranes (46% vs. 25% of total). We argue a physical-chemical case for laterally separated domains of very long chain sphingomyelins on the exoplasmic leaflet of the canalicular membrane. We bolster our hypothesis by the likelihood that the least hydrophobic, e.g., 16:0 sphingomyelin molecular species, are miscible with biliary phosphatidylcholines, and are secreted into bile. Laterally separated domains of very long chain sphingomyelins on the exoplasmic leaflet of the canalicular membrane could provide a means of sequestering cholesterol molecules prior to secretion into bile.

Animals↗
PubMed1984

Mammalian DNA enriched for replication origins is enriched for snap-back sequences.

Using the instability of replication loops as a method for the isolation of double-stranded nascent DNA, extruded DNA enriched for replication origins was obtained and denatured. Snap-back DNA, single-stranded DNA with inverted repeats (palindromic sequences), reassociates rapidly into stem-loop structures with zero-order kinetics when conditions are changed from denaturing to renaturing, and can be assayed by chromatography on hydroxyapatite. Origin-enriched nascent DNA strands from mouse, rat and monkey cells growing either synchronously or asynchronously were purified and assayed for the presence of snap-back sequences. The results show that origin-enriched DNA is also enriched for snap-back sequences, implying that some origins for mammalian DNA replication contain or lie near palindromic sequences.

Animals↗