Light microscopy techniques for live cell imaging

Since the earliest examination of cellular structures, biologists have been fascinated by observing cells using light microscopy. The advent of fluorescent labeling technologies plus the plethora of sophisticated light microscope techniques now available make studying dynamic processes in living cells almost commonplace. For anyone new to this area, however, it can be daunting to decide which techniques or equipment to try. Here, we aim to give a brief overview of the main approaches to live cell imaging, with some mention of their pros and cons.     

Landcare in Australia: Beyond the expert farmer

The landcare program has been embraced by governments, farmer organizations and conservation groups throughout Australia as offering a model for effective community action to assist the move to more sustainable resource use. Over 2,500 landcare-type groups now operate across Australia with 65,000 members including almost 30% of the farming community. This research used surveys of landcare group activity in most Australian states, a study of the regional landcare action plan (RLAP) process in the state of Victoria, and a survey of landholders in north east Victoria to assess landcare program effectiveness, and particularly, the assumptions underlying the program logic. Information provided revealed the vast scale of community participation in landcare and the considerable amount of onground work undertaken by groups and landholders. Analysis of north east landholder surveys indicated landcare participation had a significant impact upon landholder awareness of issues, level of knowledge, and adoption of best bet practices. However, research findings also highlighted a number of concerns about the program logic and implementation, and the authors suggest it is time to adopt a revised landcare program model.     

Solutions to Environmental and Economic Problems (STEEP)

This article describes one model for organizing and mobilizing scientific resources to address the highly complex and costly problem of soil erosion in the Pacific Northwest. With a U.S. Department of Agriculture grant to the agricultural experiment stations in Washington, Oregon, and Idaho, as well as supplementary state and federal funds, STEEP awards intermediate-term (15 year) grants for research in five areas: tillage and plant management, plant design, erosion and runoff predictions, pest management, and socioeconomics of erosion control. Most of the research projects require collaboration across disciplines and, in some instances, across state boundaries. After 6 years of effort the results obtained with STEEP indicate that the model might be applicable to other regions and problems.     

Undetectable intracellular free copper: the requirement of a copper chaperone for superoxide dismutase

The copper chaperone for the superoxide dismutase (CCS) gene is necessary for expression of an active, copper-bound form of superoxide dismutase (SOD1) in vivo in spite of the high affinity of SOD1 for copper (dissociation constant = 6 fM) and the high intracellular concentrations of both SOD1 (10 microM in yeast) and copper (70 microM in yeast). In vitro studies demonstrated that purified Cu(I)-yCCS protein is sufficient for direct copper activation of apo-ySOD1 but is necessary only when the concentration of free copper ions ([Cu]free) is strictly limited. Moreover, the physiological requirement for yCCS in vivo was readily bypassed by elevated copper concentrations and abrogation of intracellular copper-scavenging systems such as the metallothioneins. This metallochaperone protein activates the target enzyme through direct insertion of the copper cofactor and apparently functions to protect the metal ion from binding to intracellular copper scavengers. These results indicate that intracellular [Cu]free is limited to less than one free copper ion per cell and suggest that a pool of free copper ions is not used in physiological activation of metalloenzymes.     

Epidermal viral immunity induced by CD8alpha+ dendritic cells but not by Langerhans cells

The classical paradigm for dendritic cell function derives from the study of Langerhans cells, which predominate within skin epidermis. After an encounter with foreign agents, Langerhans cells are thought to migrate to draining lymph nodes, where they initiate T cell priming. Contrary to this, we show here that infection of murine epidermis by herpes simplex virus did not result in the priming of virus-specific cytotoxic T lymphocytes by Langerhans cells. Rather, the priming response required a distinct CD8alpha+ dendritic cell subset. Thus, the traditional view of Langerhans cells in epidermal immunity needs to be revisited to accommodate a requirement for other dendritic cells in this response.     

Effect of collector temperature and conductivity on the chain conformation of nylon 6 electrospun nanofibers

Electrospinning is a versatile process used to prepare micro- and nano- sized fibers from various polymer solution. Here, we dealt with the variation in the morphology of nylon 6 electrospun nanofibers and their polymorphism depending on the type and physical state of the collectors. SEM study showed that the fiber diameter was increased from 80 to 103 nm while it was collected in water bath. Similarly the fiber diameter and bonding was increased 103 to 115 nm with the temperature whereas it was linearly decreased 103 to 90 nm with the conductivity of the water bath. Spectroscopic analysis (FT-Raman, FT-IR) showed that the polymorphism of nylon 6 depended on the types of collector (aluminum sheet and water bath). Nylon 6 electrospun nanofibers display theγ-phase while collected in aluminum sheet andα-phase while collection in water bath. The extent of transformation fromγ- toα-phase was linearly increased with temperature and conductivity of the water bath.     

Confirmation of quantitative trait loci using a low-density single nucleotide polymorphism map for twinning and ovulation rate on bovine chromosome 5

Traditional genetic selection in cattle for traits with low heritability, such as reproduction, has had very little success. With the addition of DNA technologies to the genetic selection toolbox for livestock, the opportunity may exist to improve reproductive efficiency more rapidly in cattle. The US Meat Animal Research Center Production Efficiency Population has 9,186 twinning and 29,571 ovulation rate records for multiple generations of animals, but a significant number of these animals do not have tissue samples available for DNA genotyping. The objectives of this study were to confirm QTL for twinning and ovulation rate previously found on BTA5 and to evaluate the ability of GenoProb to predict genotypic information in a pedigree containing 16,035 animals when using genotypes for 24 SNP from 3 data sets containing 48, 724, or 2,900 animals. Marker data for 21 microsatellites on BTA5 with 297 to 3,395 animals per marker were used in conjunction with each data set of genotyped animals. Genotypic probabilities for females were used to calculate independent variables for regressions of additive, dominance, and imprinting effects. Genotypic regressions were fitted as fixed effects in a 2-trait mixed model analysis by using multiple-trait derivative-free REML. Each SNP was analyzed individually, followed by backward selection fitting all individually significant SNP simultaneously and then removing the least significant SNP until only significant SNP were left. Five significant SNP associations were detected for twinning rate and 3 were detected for ovulation rate. Two of these SNP, 1 for each trait, were significant for imprinting. Additional modeling of paternal and maternal allelic effects confirmed the initial results of imprinting done by contrasting heterozygotes. These results are supported by comparative mapping of mouse and human imprinted genes to this region of bovine chromosome 5.     

Effects of in vitro hemodilution of canine blood on platelet function analysis using the PFA‐100

Background: The platelet function analyzer (PFA)‐100 is a point‐of‐care instrument previously evaluated in humans and dogs. In both species, artificially prolonged platelet closure time (CT) occurs with anemia. Reliability of the analyzer in dogs becomes a concern when the HCT is between 0.25 and 0.35 L/L. Objective: The objective of this study was to further define the level of HCT at which CT is prolonged, using in vitro diluted canine blood. Methods: Citrated whole blood samples were collected from 22 healthy dogs. Initial HCT was determined and autologous platelet‐rich plasma was added to samples to achieve HCTs of 0.33, 0.30, and 0.27 L/L. CT was determined in duplicate on the PFA‐100 using collagen/adenosine‐5′‐diphosphate cartridges. Results: Compared with the initial CT in samples with HCT 0.39–0.54 L/L (CT mean±SD=57.8±5.75 seconds), significantly prolonged CTs were found in hemodiluted samples with HCT 0.33 L/L (61.1±4.64 seconds), 0.30 L/L (64.3±6.79 seconds), and 0.27 L/L (70.8±7.90 seconds) (P=0.029; repeated measures ANOVA). Conclusion: Although statistical differences were found, further studies are needed to determine the clinical significance of the mild prolongation in CT associated with mild anemia. Until then, dogs with HCTs slightly <0.35 L/L should be evaluated cautiously for platelet dysfunction using the PFA‐100.     

Altered pesticide use on transgenic crops and the associated general impact from an environmental perspective

The large-scale commercial cultivation of transgenic crops has undergone a steady increase since their introduction 10 years ago. Most of these crops bear introduced traits that are of agronomic importance, such as herbicide or insect resistance. These traits are likely to impact upon the use of pesticides on these crops, as well as the pesticide market as a whole. Organizations like USDA-ERS and NCFAP monitor the changes in crop pest management associated with the adoption of transgenic crops. As part of an IUPAC project on this topic, recent data are reviewed regarding the alterations in pesticide use that have been observed in practice. Most results indicate a decrease in the amounts of active ingredients applied to transgenic crops compared with conventional crops. In addition, a generic environmental indicator -- the environmental impact quotient (EIQ) -- has been applied by these authors and others to estimate the environmental consequences of the altered pesticide use on transgenic crops. The results show that the predicted environmental impact decreases in transgenic crops. With the advent of new types of agronomic trait and crops that have been genetically modified, it is useful to take also their potential environmental impacts into account.