Microm Pore Size Research Articles

Pollution prevention and control of aqueous extract of Astragali Radix processed with ZrO2 inorganic ceramic membrane micro-filtration

To study the measures for preventing and controlling the pollution of aqueous extract of Astragali Radix proceeded with inorganic ceramic membrane micro-filtration, in order to find effective measures for preventing and controlling the membrane pollution. The resistance distribution, polymer removal and changes in physical and chemical parameters of the zirconium oxide film of different pore diameters were determined to analyze the state or location of pollutants as well as the regularity of formation. Meanwhile, recoil and ultrasonic physical measures were adopted to strengthen the membrane process, in order to explore the methods for preventing and controlling the membrane pollution. When 0.2 microm of ZrO2 micro-filtrated aqueous extract of Astragali Radix, the rate of pollution was as high as 44.9%. The hole blocking resistance and the concentration polarization resistance were the main filtration resistances, while the surface deposit resistance decreased with the increase in the membrane's hold diameter; after micro-filtration, the liquid turbidity significantly reduced, with slight changes in both pH and viscosity. The 0.2 microm ZrO2 micro-filtration membrane performed better than the 0.05 microm pore size membrane in terms of conductivity. The 0. 2 microm and 0.05 microm pore diameter membranes showed better performance in the removal of pectin. The ultrasonic measure to strengthen membranes is more suitable to this system, with a flux rate up by 41.7%. The membrane optimization process adopts appropriate measures for preventing and controlling the membrane pollution, in order to reduce the membrane pollution, recover membrane performance and increase filtration efficiency.

Inhibition of migration but stimulation of proliferation of human retinal pigment epithelial cells cultured with uniform vesicles of silicone oil

It has been hypothesized that emulsification of silicone oil might stimulate preretinal membrane formation and proliferative vitreoretinopathy. This study aimed to test the effects of vesicles of silicone oil (VSO), which were prepared by a novel membrane emulsification technique, on the migration and proliferation of human retinal pigment epithelial (hRPE) cells in vitro. VSO were produced by extrusion of the oil through a Supor membrane (Pall Life Sciences, Port Washington, NY, USA) with 0.45 microm pore size under a pressure of 0.1 MPa. hRPE cells were incubated with stained 25% VSO for 24 h to assess whether hRPE cells could phagocytize VSO. A wound-healing model was used by denuding cells from a glass slide to assess the migration of hRPE cells. The cells were then incubated with 25%, 50% and 100% VSO with or without serum for up to 72 h. The number of cells that had entered the denuded area was counted under a microscope. To assess the proliferative activity, the cells were incubated with 25%, 50% and 100% VSO, with or without serum, for 24 h. A total of 100 nuclei were examined for each slide, and the numbers of stained argyrophilic nucleolar organizer regions (AgNORs) in the nuclei were measured. The mean vesicle diameter of VSO prepared was 4.25 +/- 0.77 microm. The stained 25% VSO were phagocytized by hRPE cells. After the cells cultured with serum-free Dulbecco's Modified Eagle Medium (DMEM) were incubated with VSO, the numbers of migratory cells at higher concentrations (50% and 100%) of VSO were significantly decreased (48.9 +/- 5.37 and 10.6 +/- 3.03 respectively) compared to controls (69.9 +/- 9.88; P < 0.01). After the cells cultured with serum-free DMEM were incubated with VSO, the number of AgNORs in nucleus at 100% VSO was significantly increased compared to controls (3.1 +/- 0.72 vs 1.6 +/- 0.6; P < 0.01). The result indicated that VSO inhibited the migration but stimulated the proliferation of hRPE cells.

Simultaneous Concentration of Enterococci and Coliphage from Marine Waters using a Dual Layer Filtration System

Concentrating diverse microbes in a time and cost effective manner is an essential component in water quality monitoring of recreational beaches. Historically, detection of bacteria and viruses requires two different capture methods to detect both types of organisms in a given water sample. The purpose of this present study was to evaluate a newly devised dual layered filtration system, which was developed to simultaneously concentrate both viruses and bacteria in one step from marine waters. An apparatus was designed to accommodate two 90-mm diam., 0.45 microm pore size membranes in series, one on top of the other. The top polyvinylidene fluoride (PVDF) membrane was used to filter bacteria by physical straining while the bottom HA membrane retained viruses through adsorption. Results indicated that the dual layer filtration system recovered 83+/-14% of the test bacteria (Enterococcus fecalis) and 81+/-28% of the test virus (MS2 coliphage) on the top and bottom membranes, respectively. This research demonstrates the potential of using a dual layered filtration system for the simultaneous concentration of both bacteria and viruses on separate filters from recreational beach waters. This system is relatively simple to use, inexpensive, and has the potential to be suitable for routine monitoring. This study serves as a proof of concept for the technique. Additional experiments are needed to evaluate the system on a variety of different bacteria and viruses as well as on water with different physical and chemical parameters.

Design and performance evaluation of a medium flow sampler for airborne brominated flame retardants (BFRs)

Brominated flame retardants (BFRs), including polybrominated diphenyl ethers (PBDEs), have emerged as important and ubiquitous environmental pollutants, and there is a need to accurately measure airborne levels of these chemicals in both indoor and outdoor applications. We review and suggest performance criteria for BFR sampling systems, and then present the design of a new medium flow active sampler. The sampler uses a PTFE filter (47 mm, 1 microm pore size) in front of a polyurethane foam (PUF) adsorbent plug (22 mm dia, 76 mm length) with a nominal flow rate of 15 L min(-1) and a sampling period of one week, giving a sampling volume of 150 m(3). The sampler was evaluated using co-located systems to test precision, backup PUFs to test breakthrough, and distributed volume sampling to test linearity. Field experiments were conducted in five commercial buildings, one residence and outdoors at an urban site. A total of 20 BDE congeners were quantified. After appropriate cleaning of the PUF adsorbent, blank levels were negligible. Method detection limits (MDLs) were sufficiently low to quantify BDE congeners 17, 28, 71, 47, 100 and 99 in ambient air, and more than adequate to quantify these and other congeners in indoor air, where levels are typically much higher. The relative absolute deviation (RAD), based on distributed volume samples, ranged from 21% (BDE-71) to 81% (BDE-75) for indoor samples, and was somewhat higher for ambient samples. Only minimal breakthrough was detected in back-up samples, and over 80% of the samples had very low or negligible breakthrough. Humidity did not influence sampler performance. Overall, the medium-flow sampler can accurately measure PBDEs over a wide range of concentrations and applications.

Regulation of cathepsin X overexpression in H. pylori‐infected gastric epithelial cells and macrophages

Cathepsin X (CTSX) is strongly up-regulated in Helicobacter pylori-infected gastric mucosa and intestinal-type gastric cancer. The overexpression of CTSX is mediated predominantly by associated macrophages; depends on a functional type IV-secretion system; and leads to increased migration of gastric epithelial cells. In the present study, we analysed the role of CagA in CTSX overexpression and identified H. pylori-induced inflammatory factors and signalling pathways required for stimulating CTSX expression by H. pylori. Gastric epithelial cells were co-cultured with macrophages in Transwell chambers of 0.4 microm pore size, enabling exchange of fluids but retracting H. pylori. N87 gastric epithelial cells were infected with H. pylori P1 wild-type strain in the presence of inhibitors for p38, JNK, and ERK1/2 signal transduction pathways. Furthermore, cytokines and growth factors were tested for their regulatory function using inhibitory antibodies, and their gene expression was studied by quantitative RT-PCRs and western blots. CTSX is strongly up-regulated at both the mRNA and the protein levels by TNF-alpha, IL-1beta, IL-6, and IL-8, depending on cell type. All these cytokines were found to be increased by five- to ten-fold in macrophages by H. pylori infection of co-cultured N87 gastric epithelial cells. In macrophages, H. pylori up-regulated CTSX via ERK1/2 signalling pathways, and in N87 cells via JNK irrespective of p38 signalling. Our results suggest that H. pylori induced overexpression of CTSX in macrophages and epithelium through specific cytokines that are initiated by CagA-dependent pathways in a cell type-dependent manner.

Source Apportionment of in Vitro Reactive Oxygen Species Bioassay Activity from Atmospheric Particulate Matter

Recent atmospheric particulate matter health studies have suggested that the redox activity is an important factor in particulate matter toxicology, and that reactive oxygen species (ROS) activity may be an important characteristic of particulate matter that is associated with adverse health effects. In this study, associations between atmospheric particulate matter sources and in vitro ROS activities are investigated. Ambient concentrations of fine particle water-soluble elements and total organic and elemental carbon were measured daily in Denver for the 2003 calendar year. The data were used in a multivariate factor analysis source apportionment model, positive matrix factorization (PMF), to determine the contributions of nine sources or factors: a mobile source factor, a water soluble carbon factor, a sulfate factor, a soil dust source, an iron source, two point sources characterized by water soluble toxic metals, a pyrotechnique factor, and a platinum group metal factor. Aqueous leachates, including water soluble and colloidal components, as well as insoluble particles that pass through a 0.2 microm pore size filter, of 45 randomly selected PM samples, were assayed to quantify ROS activity using an in vitro rat alveolar macrophage assay. Results show that PM-stimulated in vitro ROS production was significantly positively correlated with the contributions from three sources: the iron source, the soil dust source and the water soluble carbon factor. The iron source accounted for the greatest fraction of the measured variability in redox activity, followed by the soil dust and the water-soluble carbon factor. Seventy-seven percent of the in vitro ROS activity was explained by a linear combination of these three source contributions.

Top‐down and bottom‐up induced shifts in bacterial abundance, production and community composition in an experimentally divided humic lake

We examined in situ abundance and activities of the major bacterial groups in the two most distinct compartments of experimentally divided Lake Grosse Fuchskuhle (Germany). The selected south-west (SW) and north-east basin (NE) differ substantially in their major chemical and biological parameters that potentially influence the dynamics and composition of microbial communities. Water from the basins were incubated in dialysis bags, which allowed for a relatively free exchange of nutrients, limiting solutes and low molecular organic matter but fully prevented exchange of organisms. To investigate the effect of top-down and bottom-up manipulations three size fractions of water samples were produced: (i) unfiltered, (ii) pre-filtered through 5.0 microm pore size membranes to remove large particles, as well as grazers and (iii) pre-filtered through 0.8 microm filters to remove all potential bacterivores. One set of dialysis bags was either incubated in acidic SW (rich in humic matter) or in almost neutral NE basin whereas a second set was transferred from the SW to the NE basin and vice versa. Our study revealed pronounced differences in growth rates among the major bacterial groups in relation to the treatments. Members of the Betaproteobacteria, in particular of the subgroup targeted by the BETA2-870 probe, were highly abundant in both basins, and most of them belonged to the Polynucleobacter necessesarius subcluster PnecC. Their specific growth rates surprisingly increased in all treatments when being transplanted into the acidic SW basin, indicating that pH and humic substances greatly affected growth of this particular group in the lake. In contrast, members of the Sphingobacteria/Flavobacteria group of the Bacteroidetes (both basins) as well as Actinobacteria (SW basin) were less abundant, especially in the presence of flagellates (< 5.0 microm treatments). However, because of their extremely low initial numbers, grazing of heterotrophic nanoflagellates mostly controlled only a small part of the bacterial production (< or = 12%).

Trace metal speciation during dry and wet weather flows in the Tama River, Japan, by using diffusive gradients in thin films (DGT)

The labile species of Ni, Cu, Zn, Cd, and Pb in the Tama River, an urban river in Tokyo, Japan, were measured using diffusive gradients in thin films (DGT) method under dry and wet weather conditions, and the results were compared with total dissolved concentrations in hourly samples collected in parallel. A total of 10 DGT deployments were made and 251 hourly samples were collected during 2 rounds of sampling, conducted between August and October, 2006. Two types of diffusive gradients in thin films (DGT) devices-DGT-RG for labile inorganic and DGT-APA for total (inorganic and organic) labile species-were applied throughout the samplings. The proportions of metals measured by DGT, compared with the dissolved metal concentrations (filtered using a membrane of 0.5 microm pore size), were 38 +/- 5% (RG) and 45 +/- 8% (APA) for Ni and 45 +/- 22% (RG) and 53 +/- 23% (APA) for Zn. No labile Cu was detected throughout the sampling; Cu was assumed to be in stable complexed forms. Labile Pb was detected in 3 out of 10 deployments only; the rest were lower than the detection limit. Dissolved and labile Cd concentrations were below the detection limits. Three rain events encountered during the sampling periods were evaluated. Rains brought considerably higher loads of metals in dissolved form, and DGT measurements indicated that labile metal loads also increased. Selected DGT measurements were compared with the WHAM 6 speciation model and found to be similar to the model-computed results.

Porous β‐tricalcium phosphate/collagen composites prepared in an alkaline condition

Bone substitute materials with natural bone-like structure are considered to be favorable for bone regeneration. In this work, porous beta-tricalcium phosphate (beta-TCP)/collagen composite consisting of bone-like microstructural units was prepared using nanosized beta-TCP particles and alkaline-disassembled collagen. The resulting composite showed a good interconnecting porous structure with approximately 90% porosity and 100 approximately 300 microm pore size. The pore walls were dense, and the combination status of collagen and nanosized beta-TCP particles demonstrated that nanosized beta-TCP particles tightly connected collagen microfibrils as a bone-like microstructural unit. MTT and alkaline phosphatase (ALP) assays showed that the porous composite had enhanced effects on cellular proliferation and activity of osteoblast compared with a control of pure collagen. It is suggested that the adoption of nanosized beta-TCP particles is a main contribution to the formation of the composite with a bone-like microstructural unit, and the unique microstructure could be a main role for the composite to have the positive influence on osteoblast cell proliferation.

Ex Vivo Whole Embryonic Kidney Culture: A Novel Method for Research in Development, Regeneration and Transplantation

Whole metanephric organ culture represents a novel investigatory approach with potential application in many aspects of research in kidney regeneration and transplantation. We report the current status of embryonic kidney culture, discussing issues such as the appropriate culture conditions and methods, histological results, values of and limitations to the different techniques used today. To optimize this system in vitro for the benefit of future studies we focused our efforts on evaluating and developing a new durable 3-dimensional organ culture system using a uniquely modified approach. Metanephric kidneys were microdissected from the embryos of timed pregnant WT C57BL/C6 mice on days 12 to 16 of gestation (embryonic days 12 to 16). Novel perfusion channels were created in the harvested embryonic kidneys before placing them in culture. Embryonic kidneys were placed on a 0.4 microm pore size Transwell membrane, cultured in base medium at a medium gas interphase and incubated at 37C with fully humidified 5% CO2. Histological and immunocytochemical analysis was performed to evaluate for signs of necrosis, and the structural integrity and functionality of organs during culture. We confirmed histologically that our organ culture system was capable of maintaining normal kidney structures significantly longer (mean 10 days) than previously reported standard protocols. Condensation and aggregation of the metanephric mesenchyma at the tips of the ureteral bud were observed, including the formation of well developed nephrons and glomeruli without evidence of necrosis. Organ maturation occurred in a developmentally appropriate centrifugal pattern and the expression of key regulatory factors was demonstrated. Our in vitro model replicates closely the in vivo processes involved in normal kidney development. We also present what is to our knowledge the first demonstration of a durable 3-dimensional kidney culture system reported in the literature. This system may represent an uncomplicated method for in vitro kidney culture that we hope will serve as an effective adjunct to research focused on signaling pathways, development and regeneration as applied to the kidney.

Quantification of the Filterability of Freshwater Bacteria through 0.45, 0.22, and 0.1 μm Pore Size Filters and Shape-Dependent Enrichment of Filterable Bacterial Communities

Micro-filtration is a standard process for sterilization in scientific research, medical, and industrial applications, and to remove particles in drinking water or wastewater treatment. It is generally assumed, and confirmed by quantifying filtration efficiency by plating, that filters with a 0.1-0.45 microm pore size can retain bacteria. In contrast to this assumption, we have regularly observed the passage of a significant fraction of natural freshwater bacterial communities through 0.45, 0.22, and 0.1 microm pore size filters. Flow cytometry and a regrowth assay were applied in the present study to quantify and cultivate filterable bacteria. Here we show for the first time a systematic quantification of their filterability, especially their ability to pass through 0.1 microm pore size filters. The filtered bacteria were subsequently able to grow on natural assimilable organic carbon (AOC) with specific growth rates up to 0.47 h(-1). We were able to enrich bacteria communities that pass preferentially through all three pore size filters at significantly increased percentages using successive filtration-regrowth cycles. In all instances, the dominant microbial populations comprised slender spirillum-shaped Hylemonella gracilis strains, suggesting shape-dependent selection during the filtration process. This quantification of the omnipresence of microfilterable bacterial in natural freshwater and their regrowth characteristics demand a change in the sterile filtration practice used in industrial and engineering applications as well as scientific research.

Investigation of the Interaction Force between Cryptosporidium parvum Oocysts and Solid Surfaces

Interaction force profiles between single Cryptosporidium parvum oocysts and positively charged, silane-coated silica particles were measured in aqueous solutions using an atomic force microscope. The oocysts were immobilized for the measurements by entrapment in Millipore polycarbonate membranes with 3 microm pore size. Experiments were performed in both NaCl and CaCl2 solutions at ionic strengths ranging from 1 to 100 mM. For both electrolytes, the decay length of the repulsive force profile was found to be nearly independent of the ionic strength and always much larger than the theoretical Debye length of the system. In addition, the magnitude of the force was found to be essentially the same for both electrolytes, suggesting that the long-range repulsive forces are primarily steric in nature. These results support the theory that the interaction force between oocysts and surfaces is controlled by an outer, weakly charged or uncharged carbohydrate layer. Measurements were also performed with oocysts that had been deactivated using either chemical (formalin) or heat treatment. The force profiles obtained with formalin-treated oocysts appear to be essentially the same as for the untreated oocysts, whereas the profiles measured with the heat-treated oocysts show a much stronger dependence on solution ionic strength. With either the heat-treated or formalin-treated oocysts, adhesion was observed much more frequently than with untreated oocysts, which is consistent with the increased deposition rate observed with treated oocysts by Kuznar and Elimelech (Kuznar, Z. A.; Elimelech, M. Langmuir 2005, 21, 710-716). These results also suggest that treated oocysts, especially ones that have been inactivated by heating, may not be good surrogates for viable oocysts in laboratory studies.

Effect of pore size on in vitro cartilage formation using chitosan‐based hyaluronic acid hybrid polymer fibers

In this study, we successfully developed three-dimensional scaffolds fabricated from the chitosan-based hyaluronic acid hybrid polymer fibers, which can control the porous structure. To determine the adequate pore size for enhancing the chondrogenesis of cultured cells, we compared the behaviors of rabbit chondrocytes in scaffolds comprising different pore sizes (100, 200, and 400 microm pore size). Regarding the cell proliferation, there was no significant difference among the three groups. On the other hand, glycosaminoglycan contents in the 400 microm group significantly increased during the culture period, compared with those in the other groups. The ratio of type II to type I collagen mRNA level was also significantly higher in the 400 microm group than in the other groups. These results indicate that our scaffold with 400 microm pore size significantly enhances the extracellular matrix synthesis by chondrocytes. Additionally, the current scaffolds showed high mechanical properties, compared with liquid and gel materials. The data derived from this study suggest great promise for the future of a novel fabricated material with relatively large pore size as a scaffold for cartilage regeneration. The biological and mechanical advantages presented here will make it possible to apply our scaffold to relatively wide cartilaginous lesions.

Comparison of two radio-frequency plasma sterilization processes using microspot evaluation of microbial inactivation

In this study, we evaluated gas plasma surface sterilization methods in a specific sterilizer. We have introduced a new monitoring method using 0.4 microm pore size membranes, which in this study gave the information corresponding to 3000 exposed biological indicators per treatment cycle. This enabled us to compare the fraction of inoculates that showed no growth after exposure for 30 different locations in the chamber, and hereby identify weak and strong spots in the chamber with regard to sporicidal effect. Membranes were also used to expose a broad spectrum of soil bacteria for plasma treatment at four different conditions. The organisms were identified using PCR and sequencing. The test showed that Bacillus stearothermophilus spores were inactivated at the slowest rate among the tested microorganisms. Further alpha-proteobacteria (Gram negative) seemed more sensitive than the rest of the tested organisms. The microspot evaluation approach has been a most useful tool in the assessment of sterilization performance in sterilizers that do not have clear measurable parameters related to the sterilization.

Interaction Force Profiles between Cryptosporidium parvum Oocysts and Silica Surfaces

The interaction force profile between single Cryptosporidium parvum oocysts and silica particles was measured in aqueous solutions using an atomic force microscope. The oocysts were immobilized during the measurements by entrapment in Millipore polycarbonate membranes with a 3 microm pore size. Experiments were performed in both NaCl and CaCl2 solutions at ionic strengths ranging from 1 to 100 mM. For both electrolytes the decay length of the repulsive force profile, obtained via the slope of a plot of the logarithm of interaction force versus separation, was found to be essentially independent of the ionic strength and always much larger than the theoretical Debye length of the system. In addition, the magnitude of the force was found to be essentially the same for both electrolytes, suggesting that the long-range repulsive forces are primarily steric in nature. Fitting the force to an expression for the steric repulsive force between two grafted brush layers yields a layer thickness of approximately 115 nm. These results support the idea that the oocysts are covered by a relatively thick layer of uncharged (or weakly charged) carbohydrates, possibly mixed with a thinner layer of charged protein.

Effect of restricted forage intake on ruminal disappearance of bromegrass hay and a blood meal, feather meal, and fish meal supplement1

Two experiments were conducted to determine in situ disappearance of bromegrass hay and a ruminally undegraded protein (RUP) supplement in beef cattle fed restricted amounts of forage. Six Angus crossbred cattle (BW = 589 +/- 44.4 kg; three steers and three heifers) fitted with ruminal cannulas were fed chopped (2.54 cm) bromegrass hay (8.9% CP) at one of three percentages of maintenance intake (30, 55, or 80%; one steer and one heifer per treatment). In both experiments, the cattle were allowed 7 d for diet adaptation followed by 3 d of sample collection. In Exp 1, in situ bags (50 microm pore size) containing 4.1 g of brome-grass hay (OM basis) were inserted into the rumen and subsequently removed at 3, 6, 9, 12, 15, 18, 24, 36, and 48 h after insertion. Nonlinear regression models were used to determine the rapidly solubilized protein Fraction A, the potentially ruminal degradable protein Fraction B, the ruminally undegraded protein Fraction C, and protein degradation rate. Intake level did not affect (P = 0.15 to 0.95) forage protein remaining after in situ incubation or Fractions A, B, and C; however, effective ruminal degradation of hay protein tended to increase quadratically (P = 0.12) as forage intake increased. In Exp 2, 4.2 g (OM basis) of an RUP supplement (6.8% porcine blood meal, 24.5% hydrolyzed feather meal, and 68.7% menhaden fish meal) formulated to provide equal amounts of metabolizable protein across all levels of hay consumption was evaluated in a similar manner as in Exp 1. The undegraded protein fraction of the supplement did not differ (P = 0.16 to 0.74) across treatments at 3, 6, 9, and 18 h; however, increasing forage intake resulted in a linear increase (P < or = 0.06) in undegraded protein remaining at 12, 15, 24, 36, and 48 h. Dietary treatment had no affect (P = 0.30) on protein Fractions A, B, or C; however, protein degradation rate of the supplement decreased linearly (P = 0.03) as forage intake increased. Therefore, effective ruminal degradation of the supplement decreased linearly (P = 0.01) from 50.8 to 40.9% as forage intake increased from 30 to 80% of maintenance. Corresponding estimates of supplement RUP were 49.2, 56.5, and 59.1% for the 30, 55, and 80% of maintenance intake treatments, respectively. Restricting dietary intake can decrease the quantity of dietary protein that escapes ruminal degradation. Tabular estimates of RUP may not be appropriate for formulating diets to balance metabolizable protein in beef cattle consuming limited quantities of forage.

Detection of E. coli in beach water within 1 hour using immunomagnetic separation and ATP bioluminescence.

The contamination of beach waters occurs from the discharge of storm water and sanitary sewer overflows containing faecal material. Additional faecal material derives from discharge of animals and waterfowl. In order to protect public from exposure to faecal-contaminated water, it is required to test enteric indicators in beach water. The problem is that the traditional culture-based methods cannot meet this goal because it takes too long (>24 h), so the results are not available until a day later. A rapid method for testing beach water for Escherichia coli within 1 h has been developed. Immunomagnetic separation (IMS) and ATP bioluminescence were used for selective capture and quantification, respectively. This rapid method was compared to the current method (m-TEC) using beach water samples. The beach samples were prefiltered with a 20 microm pore size filter in order to remove algae, plant debris and large particles. The results showed that the prefiltration step did not trap the bacteria which were present in the original water samples. The prefiltered water was then passed through a 0.45 microm pore size filter for concentration. The deposited bacteria were resuspended and then mixed with superparamagnetic polystyrene beads (diameter of 0.6 microm) that were coated with E. coli antibodies. After IMS, the quantification of the E. coli was done by ATP bioluminescence. The results obtained with IMS-ATP bioluminescence correlated well with the plate count method (Rsq = 0.93). The detection limit of the assay was about 20 CFU/100 mL, which is well below the US EPA limits for recreational water. The entire procedure can be completed in less than 1 hour. The necessary equipment is portable and was tested on-site.

Preparation and culture of hepatocyte on gelatin microcarriers.

Porous gelatin microcarriers having a diameter of 80-100 microm were prepared by the suspension method using toluene as the oil phase. Rat hepatocytes were cultured on gelatin and cytodexIII microcarriers. The cells retained its spherical shape, which is similar in vivo, and showed no morphological changes to the flat state. Hepatocyte aggregates on microcarriers maintained higher metabolic functions than monolayer cells. Pore size of microcarrier plays an important role in the attachment and metabolic function of cells in culture. Phase-contrast micrograph and cell activity showed that hepatocytes cultures on gelatin microcarrier of <1 microm pore size is better than that of 5-10 microm.

Anti-inflammatory effect of rutin on rat paw oedema, and on neutrophils chemotaxis and degranulation

Rutin, a natural flavone derivative, is known for its pharmacological properties. We have previously reported that this flavonol exerted a potent inhibitory effect on respiratory burst of fMet-Leu-Phe-stimulated neutrophils, as well as on phosphoinositide 3-kinase gamma activity in a cell free system. In the present study, the anti-inflammatory effect of rutin was investigated in vivo and in vitro. rutin or aspirin (100 mg/kg, body weight) were given orally to rats 1 hour before paw oedema induction, using lambda-carrageenan 1%. The rat paw volume was measured by mean of plethysmometer, initially and during 6 hours. The chemotaxis of neutrophils towards 10(-7) M fMet-Leu-Phe was performed using 48-well chemotaxis chamber. Neutrophils that migrated through 5 microm pore size polycarbonate filter, in presence or in absence of rutin, were counted microscopically. Elastase exocytosis of either phorbol 12-myristate 13-acetate or fMet-Leu-Phe/cytochalasin B-stimulated neutrophils was assessed in absence or in presence of rutin using the synthetic substrate N-Suc-Ala-Ala-Ala-p-nitroanilide. The absorbance of released p-nitroaniline was measured at 405 nm using microplate reader. The maximal swelling in placebo group was observed at 5 hours, after lambda-carrageenan injection. Oral administration of rutin reduced rat paw swelling starting 2 hours after lambda-carrageenan injection. Rutin reduced significantly (p < 0.05) and in a dose-dependant manner the polymorphonuclear neutrophils chemotaxis to fMet-Leu-Phe. Furthermore, elastase exocytosis, induced by both stimuli, was partially inhibited by rutin up to 25 microM. The present study revealed that rutin possesses anti-inflammatory properties.

Porous PEOT/PBT scaffolds for bone tissue engineering: preparation, characterization, and in vitro bone marrow cell culturing.

The preparation, characterization, and in vitro bone marrow cell culturing on porous PEOT/PBT copolymer scaffolds are described. These scaffolds are meant for use in bone tissue engineering. Previous research has shown that PEOT/PBT copolymers showed in vivo degradation, calcification, and bone bonding. Despite this, several of these copolymers do not support bone marrow cell growth in vitro. Surface modification, such as gas-plasma treatment, is needed to improve the in vitro cell attachment. Porous structures were prepared using a freeze-drying and a salt-leaching technique, the latter one resulting in highly porous interconnected structures of large pore size. Gas-plasma treatment with CO(2) generated a surface throughout the entire structure that enabled bone marrow cells to attach. The amount of DNA was determined as a measure for the amount of cells present on the scaffolds. No significant effect of pore size on the amount of DNA present was seen for scaffolds with pore sizes between 250-1000 microm. Light microscopy data showed cells in the center of the scaffolds, more cells were observed in the scaffolds of 425-500 microm and 500-710 microm pore size compared to the ones with 250-425 microm and 710-1000 microm pores.