Microfluidic analysis of antibody specificity in a compact disk format

A new and flexible technology for high throughput analysis of antibody specificity and affinity is presented. The method is based on microfluidics and takes advantage of compact disks (CDs) in which the centrifugal force moves fluids through microstructures containing immobilized metal affinity chromatography columns. Analyses are performed as a sandwich assay, where antigen is captured to the column via a genetically attached His6-tag. The antibodies to be analyzed are applied onto the columns. Thereafter, fluorescently labeled secondary antibodies recognize the bound primary antibodies, and detection is carried out by laser-induced fluorescence. The CDs contain 104 microstructures enabling analysis of antibodies against more than 100 different proteins using a single CD. Importantly, through the three-dimensional visualization of the binding patterns in a column it is possible to separate high affinity from low affinity binding. The method presented here is shown to be very sensitive, flexible and reproducible.     

PEDF: a potential molecular therapeutic target with multiple anti-cancer activities

Pigment epithelium-derived factor (PEDF) is an endogenously produced protein that is widely expressed throughout the human body, and exhibits multiple and varied biological activities. Already established as a potent anti-angiogenic molecule, PEDF has recently shown promise as a potential anti-tumour agent, causing both direct and indirect tumour suppression. Here, we explore the unique anti-tumour properties of PEDF and discuss its role as an effective anti-angiogenic, anti-proliferative and pro-differentiation factor. We also discuss the prospects for PEDF therapy and the need for a closer evaluation of issues such as delivery, stability and potential toxicity.     

Functional characterization of the powdery mildew susceptibility gene <Emphasis Type="Italic">SmMLO1</Emphasis> in eggplant (<Emphasis Type="Italic">Solanum melongena</Emphasis> L.)

Eggplant (Solanum melongena L.) is one of the most important vegetables among the Solanaceae and can be a host to fungal species causing powdery mildew (PM) disease. Specific homologs of the plant Mildew Locus O (MLO) gene family are PM susceptibility factors, as their loss of function results in a recessive form of resistance known as mlo resistance. In a previous work, we isolated the eggplant MLO homolog SmMLO1. SmMLO1 is closely related to MLO susceptibility genes characterized in other plant species. However, it displays a peculiar non-synonymous substitution that leads to a T → M amino acid change at protein position 422, in correspondence of the MLO calmodulin-binding domain. In this study, we performed the functional characterization of SmMLO1. Transgenic overexpression of SmMLO1 in a tomato mlo mutant compromised resistance to the tomato PM pathogen Oidium neolycopersici, thus indicating that SmMLO1 is a PM susceptibility factor in eggplant. PM susceptibility was also restored by the transgenic expression of a synthetic gene, named s-SmMLO1, encoding a protein identical to SmMLO1, except for the presence of T at position 422. This indicates that the T → M polymorphism does not affect the protein role as PM susceptibility factor. Overall, the results of this work are of interest for the functional characterization of MLO proteins and the introduction of PM resistance in eggplant using reverse genetics.     

Risk screening of non-native and translocated freshwater fish species in a Mediterranean-type shallow lake: Lake Marmara (West Anatolia)

Risk screening tools to identify species with a high or low risk of invasiveness are being increasingly used for effective management purposes. Amongst the available tools, the Fish Invasiveness Screening Kit (FISK) has been used extensively and successfully in large risk assessment (RA) areas, and was recently upgraded to the new generic tool Aquatic Species Invasiveness Screening Kit (AS-ISK). The aim of the present study was to assess with AS-ISK the invasive potential of introduced non-native and translocated fishes in a Mediterranean-type shallow lake (Lake Marmara) located in west Anatolia (Turkey). Based on independent evaluations of 35 species by two assessors, calibration of AS-ISK resulted in a threshold score of -3.65, which reliably distinguished between potentially invasive (high risk) and potentially non-invasive (medium to low risk) fishes. Of the 35 species assessed, 17 were categorised as ‘low risk’ and included native/endemic and translocated natives, and the remaining 18 as ‘high risk’ and comprised non-natives and translocated natives. Carassius gibelio had the highest score in the Climate Change Assessment section, suggesting that it might potentially impact on the native fish fauna under likely climate change scenarios for the RA area. Some cool water non-native (Oncorhynchus mykiss, Salvelinus fontinalis), translocated native (Luciobarbus lydianus) and endemic species (Ladigesocypris mermere) in the catchment will likely be affected negatively by predicted climate change conditions.     

Size‐based interactions across trophic levels in food webs of shallow Mediterranean lakes

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Identification of rosmarinic acid and sulfated flavonoids as inhibitors of microfouling on the surface of eelgrass Zostera marina

A bioassay-guided approach was used to identify defense compounds that are present on the surface of Zostera marina and which inhibit settlement of microfoulers at natural concentrations. Moderately polar eelgrass surface extracts inhibited the settlement of seven marine bacteria and one yeast that originated from non-living substrata. In contrast, five other bacterial strains that had been directly isolated from eelgrass surfaces were all insensitive, which suggested a selective effect of surface metabolites on the microbial communities present on eelgrass. Bioassay-guided isolation of active compounds from the extracts in combination with UPLC-MS and ¹H-NMR spectroscopy resulted in the identification of rosmarinic acid, luteolin-7-sulfate and diosmetin-7-sulfate or its isomer chrysoeriol-7-sulfate. All three compounds are nontoxic repellents, as they did not inhibit bacterial growth, but prevented bacterial settlement in a dose-dependent manner. Between 15.6 and 106.8 μg ml^?1 of rosmarinic acid were present on the eelgrass surface, enough for half maximal settlement inhibition of bacteria.     

Sustainability assessment and comparison of efficacy of four P-inactivation agents for managing internal phosphorus loads in lakes: sediment incubations

A novel application of a continuous flow incubation system (CFIS) was used to assess four phosphorus (P) inactivation agents—alum, Phoslock™, a new modified zeolite (Z2G1 or Aqual-P™), and allophone—when used as sediment capping agents to manage internal P loads in lakes. The CFIS technique allowed combined efficacy and sustainability assessment, including: (1) flux measurements during simulation of stratified (anoxic) and mixed (aerobic) conditions on the same sediment through multiple cycles to assess the longevity of a range of product doses; (2) simulation of a summer algal bloom collapse and subsequent burial of the products; and (3) investigation of non-target effects on nitrification and denitrification processes at the sediment–water interface. Minimum P-removal dose rates were found to differ substantially at 80 g m⁻² for alum, 190 g m⁻² for Z2G1, 220 g m⁻² for allophane and 280 g m⁻² for Phoslock™, for similar capping layer thickness of about 2 mm, and would be effective for at least 4 years. All products temporarily suppressed nitrification and denitrification under aerobic conditions, and it may be important to minimise product application to any permanently aerobic zones, such as the littoral areas of a lake. While the aluminium (Al)-based products did not enhance Al fluxes in the CFIS, lanthanum (La) was released at a near constant rate of around 2 mg La m⁻² day⁻¹ from the Phoslock™ treatments over a period of at least 14 days. Spatial variability of sediment P, bioturbation, and burial are factors that will affect up-scaling these results to a whole lake.