Prevention of bridge binding in immunoassays: a general estradiol tracer structure

Iodinated estradiol tracers were synthesized with three different bridges connecting the radiolabelled moiety to the steroid core: Hemisuccinate, carboxymethyloxime and amide. Taking these iodinated tracers in combination with ten antibodies raised against estradiol-6-CMO-albumin, titers and slopes of calibration curves have been compared to the corresponding data using a 3H tracer. The data indicate that the tracer with the amide bridge is recognized similarly to the tritiated estradiol by all antibodies tested, whereas the two other iodinated tracers exhibit substantial bridge binding. The results suggest that the amide tracer structure can generally be used to improve the quality of estradiol antibodies suffering from bridge binding effects.     

The use of lead as an ionic tracer for investigating routes of passive fluid transfer

Several inorganic ions, including lead, barium, silver, and thallium, have been tested as possible tracers for demonstrating fluid-accessible channels in functional epithelia at the ultrastructural level. The most useful of the ionic tracers examined was the lead (plumbous) ion, administered for short time intervals (less than 2 min) and "captured" with phosphate used as the buffer in the fixative. Passive fluid and ion-accessible channels of rat parotid salivary gland have been examined with this method. At short tracer infusion times (0.5-1.0 min), localization of the tracer was primarily extracellular, although intracellular deposits were observed in the following sites: smooth membrane-delimited endocytic vesicles of both epithelial and connective tissue cells, inner Golgi cisternae, and occasional cisternae of rough endoplastic reticulum. The lead tracer readily penetrated tight junctions between parotid acinar cells but rarely passed through the tight junctions between intercalated duct cells and did not penetrate junctions of striated duct cells. Fat cells observed in the stroma of this gland were the only cells that exhibited lead tracer in the cytosol, suggesting that the plasmalemma of this cell type is more permeable to exogenous ions than the plasmalemma of other cell types present in this gland.     

Stable isotopes of minerals as metabolic tracers in human nutrition research

Enriched stable isotopes used as tracers have proven to be valuable in studies of the absorption and metabolism of minerals. Unlike radioisotopes, they can be used in high-risk population groups such as infants, children, and pregnant or lactating women. Estimates of mineral absorption can be made from the oral administration of a single tracer or from two tracers, one given orally and the other intravenously (IV). It is possible to determine the metabolism of the mineral with modeling based on the amount of the tracer or tracers in different biological samples. One of the key decisions in studies of this type is determining which enriched isotope and what amount to use. An example is given of calculations to estimate and compare the amounts of tracers needed for an absorption study. Methods for calculating the amounts of tracer in oral and IV doses are presented, and limits of detection and quantitation are discussed in terms of percent of enrichment and related to isotope ratio measurement precision. A general review of the use of mass spectrometric instruments for quantifying various stable isotopes is given.     

Rational design of 13C-labeling experiments for metabolic flux analysis in mammalian cells

ABSTRACT: BACKGROUND: 13C-Metabolic flux analysis (13C-MFA) is a standard technique to probe cellular metabolism and elucidate in vivo metabolic fluxes. 13C-Tracer selection is an important step in conducting 13C-MFA, however, current methods are restricted to trial-and-error approaches, which commonly focus on an arbitrary subset of the tracer design space. To systematically probe the complete tracer design space, especially for complex systems such as mammalian cells, there is a pressing need for new rational approaches to identify optimal tracers. RESULTS: Recently, we introduced a new framework for optimal 13C-tracer design based on elementary metabolite units (EMU) decomposition, in which a measured metabolite is decomposed into a linear combination of so-called EMU basis vectors. In this contribution, we applied the EMU method to a realistic network model of mammalian metabolism with lactate as the measured metabolite. The method was used to select optimal tracers for the two free fluxes in the system, the oxidative pentose phosphate pathway (oxPPP) flux and anaplerosis by pyruvate carboxylase (PC). Our approach was based on sensitivity analysis of EMU basis vector coefficients with respect to free fluxes. Through efficient grouping of coefficient sensitivities, simple tracer selection rules were derived for high-resolution quantification of the fluxes in the mammalian network model. The approach resulted in a significant reduction of the number of possible tracers and the feasible tracers were evaluated using numerical simulations. Two optimal, novel tracers were identified that have not been previously considered for 13C-MFA of mammalian cells, specifically [2,3,4,5,6-13C]glucose for elucidating oxPPP flux and [3,4-13C]glucose for elucidating PC flux. We demonstrate that 13C-glutamine tracers perform poorly in this system in comparison to the optimal glucose tracers. CONCLUSIONS: In this work, we have demonstrated that optimal tracer design does not need to be a pure simulation-based trial-and-error process; rather, rational insights into tracer design can be gained through the application of the EMU basis vector methodology. Using this approach, rational labeling rules can be established a priori to guide the selection of optimal 13C-tracers for high-resolution flux elucidation in complex metabolic network models.     

Determination of stable molybdenum isotopes by thermionic mass spectrometry for tracer studies of molybdenum plant metabolism

A tracer technique based on multiple stable Mo isotopes and thermionic quadrupole mass spectrometry for isotopic analysis of plant tissue was developed and has been applied in the long-term study of foliar absorption of Mo by potato plants. As several tracers have been used the multivariate linear regression methods has been applied to calculate the portions of tracer Mo present in the potato samples from the isotopic ratios measured and to estimate their reproducibilities.In this paper solely the tracer portions transferred from the leaf into the tuber of the potato have been determined. Dependent on the time of contamination, tracer portions of 0.2 to 12% have been measured, corresponding to a maximum of 2.3% of the foliar application of the tracer molybdate. The reproducibilities of the tracer method as a whole (analytical determination and calculation of the tracer portions in the tracer plant samples) amounted to 7% at the maximum (with one exception); by contrast, the individual differences between the three plants investigated were much larger (up to 80%).     

Measurement of human mixed muscle protein fractional synthesis rate depends on the choice of amino acid tracer

The goal of this study was to discover whether using different tracers affects the measured rate of muscle protein synthesis in human muscle. We therefore measured the mixed muscle protein fractional synthesis rate (FSR) in the quadriceps of older adults during basal, postabsorptive conditions and mixed meal feeding (70 mg protein x kg fat-free mass(-1) x h(-1) x 2.5 h) by simultaneous intravenous infusions of [5,5,5-(2)H(3)]leucine and either [ring-(13)C(6)]phenylalanine or [ring-(2)H(5)]phenylalanine and analysis of muscle tissue samples by gas chromatography-mass spectrometry. Both the basal FSR and the FSR during feeding were approximately 20% greater (P < 0.001) when calculated from the leucine labeling in muscle tissue fluid and proteins (fasted: 0.063 +/- 0.005%/h; fed: 0.080 +/- 0.007%/h) than when calculated from the phenylalanine enrichment data (0.051 +/- 0.004 and 0.066 +/- 0.005%/h, respectively). The feeding-induced increase in the FSR ( approximately 20%; P = 0.011) was not different with leucine and phenylalanine tracers (P = 0.69). Furthermore, the difference between the leucine- and phenylalanine-derived FSRs was independent of the phenylalanine isotopomer used (P = 0.92). We conclude that when using stable isotope-labeled tracers and the classic precursor product model to measure the rate of muscle protein synthesis, absolute rates of muscle protein FSR differ significantly depending on the tracer amino acid used; however, the anabolic response to feeding is independent of the tracer used. Thus different precursor amino acid tracers cannot be used interchangeably for the evaluation of muscle protein synthesis, and data from studies using different tracer amino acids can be compared qualitatively but not quantitatively.     

以标志物对长江上游漂流性鱼卵漂流方式的研究

2008年4月7日在长江上游重庆江津-珞璜31km的江段采用标志物对自然条件下大流量河流漂流性鱼卵的漂流方式进行模拟实验。结果显示在流量稳定的情况下, 在同一投放点投放的标志物的平均漂流速度具有显著差异, 标志物在河流中的漂流密度以高峰值和低峰值相间的方式出现, 高峰值随漂流时间的增加呈逐渐下降趋势。在水平方向上标志物的分布以主流水域的漂流密度高于缓流水域, 缓流水域未采集到标志物。采用传统产卵规模计算方法对标志物投放数量进行估算,各组的估算数量均小于实际投放数量, 估算数量的准确性随漂流时间的增加而降低, 总体估算数量为实际投放数量的16.6%。根据实验结果, 建议在长江上游鱼类产卵规模计算中引入时间系数以弥补随漂流时间的增加而引起的估算准确性降低。       

The problem of estimating wind drift in migrating birds

Whether migrating birds compensate for wind drift or not is a fundamental question in bird migration research. The procedures to demonstrate and quantitatively estimate wind drift or compensation are fraught with difficulties and pitfalls. In this paper, we evaluate four methods that have been used in several studies over the past decades. We evaluate the methods by analysing a model migratory movement with a realistic scatter in flight directions, for the ideal cases of full drift and complete compensation. Results obtained with the different methods are then compared with the "true behaviour" of the model movement, illustrating that spurious patterns of drift and compensation arise in some cases. We also illustrate and evaluate the different methods of estimating drift for a real case, based on tracking radar measurements of bird migration in relation to winds. Calculating the linear regression of mean geographic track (resulting flight direction) and heading directions (directions of the birds' body axis) of a migratory movement under different wind conditions in relation to the angle alpha (the angle between mean track and heading) always provides robust and reliable results. Comparing mean flight directions between occasions with winds from the left and right of the mean flight direction of the whole migratory movement also always provides expected and correct measures of drift. In contrast, regressions of individual flight directions in relation to alpha (the angle between track and heading for the specific individuals or flocks) are liable to produce biased and spurious results, overestimating compensation/overcompensation if following winds dominate in the analysis and overestimating drift/overdrift if opposed winds are dominating. Comparing mean directions for cases with winds from the left and right in relation to individual flight directions also gives biased and spurious results unless there is full variation in wind directions or an equal distribution of crosswinds from left and right. The results of the methodological evaluation and the analysis of the real case indicate that some earlier analyses of wind drift may have to be re-evaluated.     

Tracing root permeability: comparison of tracer methods

Root epidermis and apoplastic barriers (endodermis and exodermis) are the critical root structures involved in setting up plant-soil interface by regulating free apoplastic movement of solutes within root tissues. Probing root apoplast permeability with “apoplastic tracers” presents one of scarce tools available for detection of “apoplastic leakage” sites and evaluation of their role in overall root uptake of water, nutrients, or pollutants. Although the tracers are used for many decades, there is still not an ideal apoplastic tracer and flawless procedure with straightforward interpretation. In this article, we present our experience with the most frequently used tracers representing various types of chemicals with different characteristics. We examine their behaviour, characteristics, and limitations. Here, we show that results gained with an apoplastic tracer assay technique are reliable but depend on many parameters–chemical properties of a selected tracer, plant species, cell wall properties, exposure time, or sample processing.     

Odour sampling. 2. Comparison of physical and aerodynamic characteristics of sampling devices: a review

Sampling devices differing greatly in shape, size and operating condition have been used to collect air samples to determine rates of emission of volatile substances, including odour. However, physical chemistry principles, in particular the partitioning of volatile substances between two phases as explained by Henrys Law and the relationship between wind velocity and emission rate, suggests that different devices cannot be expected to provide equivalent emission rate estimates. Thus several problems are associated with the use of static and dynamic emission chambers, but the more turbulent devices such as wind tunnels do not appear to be subject to these problems. In general, the ability to relate emission rate estimates obtained from wind tunnel measurements to those derived from device-independent techniques supports the use of wind tunnels to determine emission rates that can be used as input data for dispersion models.     

Wind tunnel as a tool in bird migration research

Wind tunnels, in which birds fly against an artificially generated air flow, have since long been used to evaluate aerodynamic properties of steady bird flight. A new generation of wind tunnels has also allowed the many processes associated with migratory flights to be studied in captivity. We review how wind tunnel studies of aerodynamics and migratory performance together have helped advancing our understanding of bird migration. Current migration theory is based on the power‐speed relationship of flight as well as flight range equations, both of which can be evaluated using birds flying in wind tunnels. In addition, and depending on wind tunnel properties, performance during gliding and climbing flight, and effects of air pressure, humidity and turbulence on bird flight has been measured. Long‐distance migrant species have been flown repeatedly for up to 16 h non‐stop, allowing detailed studies of the energy expenditure, fuel composition, protein turnover, water balance, immunocompetence and stress associated with sustained migratory flights. In addition, wind tunnels allow the fuelling periods between migratory flights to be studied from new angles. We end our review by suggesting several important topics for future wind tunnel studies, ranging from on of the key questions remaining, the efficiency at which chemical power in converted to mechanical power, to new useful avenues, such as improving and calibrating the techniques used for tracking of individual birds in the wild.     

Drift studies--comparison of field and wind tunnel experiments

Drift at pesticide application leads to a pollution of non-target crops, non-target species and surface water. Spray drift is influenced by many factors like environmental conditions, vegetation, technical conditions, and physical properties of the tank mixes and influenced by Chemicals. Field experiments to characterise spray drift effects with the risk of permanent changing weather conditions can be supported by wind tunnel experiments. Wind tunnel experiments do not lead to the same soil deposition curves like field experiments, but the ratio of drift reduction potential is comparable.     

风洞技术在昆虫化学通讯研究中的应用

综述了昆虫化学生态学中所使用的风洞技术。内容包括风洞的构造 ,风洞的工作条件 ,风洞操作中的技巧和数据采集 ,昆虫在风洞中表现出的典型行为特征以及一些需要注意的问题     

Assessment of the permeability properties of cryopreservation outer bags used in NHSBT

This study was carried out to investigate leakage/transport across the bag material of six outer cryopreservation bags in common use within NHS Blood and Transplant. In order to do this two different leak testing procedures; coloured dye and hydrogen tracer gas, were used. The data obtained show that a coloured dye cannot permeate through the materials both at room temperature and following storage at liquid nitrogen temperature (? 196 °C). In addition, when filled with the smallest elemental molecule, hydrogen, in the form of a tracer gas, all of the bags only allowed trace amounts of hydrogen to escape, either through the seal or the bag material. The data indicated that each of the bag materials tested would be capable of preventing bacterial or viral cross-contamination as long as the material remained intact.     

Tracing of sensory neurones and spinal motoneurones of the pigeon by injection of fluorescent dyes into peripheral nerves

The projection of peripheral sensory and motor nerves was investigated in the pigeon (Columba livia) by means of retrogradely transported fluorescent dyes. Two combinations of fluorescent tracers were used that could be identified within the same cell when excited by light of 405 nm: 1) Propidium iodide and Bisbenzimide, which label the cytoplasm orange and the nucleus blue, respectively; 2) Fast Blue, which labels the cytoplasm blue, and Nuclear Yellow, which labels the nucleus (especially the nucleolar ring) yellow. The presence of the tracers in a given cell was confirmed microspectrophotometrically. Following injection of the tracers into peripheral nerves, labelled sensory neurones were seen in the dorsal root ganglia and motoneurones of the spinal cord. The peroneal and tibial nerves projected to L2-L5 and L2-L7, respectively, whereas the median and ulnar nerves projected to C12-Th2 and C13-Th1. Double-labelled sensory neurones were observed when both peroneal and tibial, or median and ulnar nerves were injected with different tracers. This indicates that some sensory neurones possess peripheral processes that dichotomize to pass down two different peripheral nerves. Double labelling was never seen in motoneurones, or in sensory neurones after tracer injection into the sciatic and femoral nerves.     

Spray drift as affected by meteorological conditions

Spray drift can be defined as the quantity of plant protection product that is carried out of the sprayed (treated) area by the action of air currents during the application process. This continues to be a major problem in applying agricultural pesticides. The purpose of this research is to measure and compare the amount of drift for different climatological conditions under field conditions. Spray drift was determined by sampling in a defined downwind area at different positions in a flat meadow using horizontal drift collectors (sedimenting spray drift) and pipe cleaners (airborne spray drift) for a reference spraying. Meteorological conditions were monitored during each experiment. A drift prediction equation for the reference spraying was set up to predict the expected magnitude of sedimenting drift at various drift distances and atmospheric conditions (wind speed and temperature). This equation can be used to compare measurements using other spraying techniques under different weather conditions to the reference spraying. In 2005, more measurements will be performed to validate the statements and the model reflected in this paper.     

Trade‐Offs in Thin Film Solar Cells with Layered Chalcostibite Photovoltaic Absorbers

Discovery of novel semiconducting materials is needed for solar energy conversion and other optoelectronic applications. However, emerging low‐dimensional solar absorbers often have unconventional crystal structures and unusual combinations of optical absorption and electrical transport properties, which considerably slows down the research and development progress. Here, the effect of stronger absorption and weaker carrier collection of 2D‐like absorber materials are studied using a high‐throughput combinatorial experimental approach, complemented by advanced characterization and computations. It is found that the photoexcited charge carrier collection in CuSbSe₂ solar cells is enhanced by drift in an electric field, addressing a different absorption/collection balance. The resulting drift solar cells efficiency is <5% due to inherent J _SC/V _OC trade‐off, suggesting that improved carrier diffusion and better contacts are needed to further increase the CuSbSe₂ performance. This study also illustrates the advantages of high‐throughput experimental methods for fast optimization of the optoelectronic devices based on emerging low‐dimensional semiconductor materials.     

In vivo rat brain opioid receptor binding of LY255582 assessed with a novel method using LC/MS/MS and the administration of three tracers simultaneously

LY255582 is a pan opioid selective receptor antagonist that has been shown to have high affinity for mu, delta, and kappa receptors in vitro. In order to better understand the in vivo opioid receptor selectivity of LY255582, we developed in vivo receptor occupancy assays in the rat for the opioid mu, kappa and delta receptors using the occupancy tracers naltrexone, GR103545 and naltriben respectively. Individual assays for each target were established and then a "triple tracer" assay was created where all three tracers were injected simultaneously, taking advantage of LC/MS/MS technology to selectively monitor brain tracer levels. This is the first report of a technique to concurrently measure receptor specific occupancy at three opioid receptors in the same animal. The opioid subtype selective antagonists cyprodime, JDTic and naltrindole were used to validate selectivity of the assay. Examination of LY255582 in dose-occupancy experiments demonstrated a relative order of potency of mu>kappa>delta, reproducing the previously reported order determined with in vitro binding.     

Oxidation of glutamine by the splanchnic bed in humans

[1,2-(13)C(2)]glutamine and [ring-(2)H(5)]phenylalanine were infused for 7 h into five postabsorptive healthy subjects on two occasions. On one occasion, the tracers were infused intravenously for 3.5 h and then by a nasogastric tube for 3.5 h. The order of infusion was reversed on the other occasion. From the plasma tracer enrichment measurements at plateau during the intravenous and nasogastric infusion periods, we determined that 27 +/- 2% of the enterally delivered phenylalanine and 64 +/- 2% of the glutamine were removed on the first pass by the splanchnic bed. Glutamine flux was 303 +/- 8 micromol. kg(-1). h(-1). Of the enterally delivered [(13)C]glutamine tracer, 73 +/- 2% was recovered as exhaled CO(2) compared with 58 +/- 1% of the intravenously infused tracer. The fraction of the enterally delivered tracer that was oxidized specifically on the first pass by the splanchnic bed was 53 +/- 2%, comprising 83% of the total tracer extracted. From the appearance of (13)C in plasma glucose, we estimated that 7 and 10% of the intravenously and nasogastrically infused glutamine tracers, respectively, were converted to glucose. The results for glutamine flux and first-pass extraction were similar to our previously reported values when a [2-(15)N]glutamine tracer [Matthews DE, Morano MA, and Campbell RG, Am J Physiol Endocrinol Metab 264: E848-E854, 1993] was used. The results of [(13)C]glutamine tracer disposal demonstrate that the major fate of enteral glutamine extraction is for oxidation and that only a minor portion is used for gluconeogenesis.     

Metabolism of high-density lipoproteins in rainbow trout.

Trout high-density lipoproteins have been labelled with residualizing tracers for the lipid and protein moieties ([3H]cholesteryloleyl ether and 125I-tyramine-cellobiose, respectively). Plasma kinetics and tissue site of catabolism were determined for both tracers. The lipid tracer was cleared about twice as fast from the blood as the protein tracer (half lifes were 63.5 and 125.3 h, respectively). This selective removal of lipid from the lipoprotein was mainly accomplished by the higher liver uptake of the cholesteryl ether. The main catabolic site for HDL protein was kidney tissue. This data established the existence of differential HDL catabolism in a lower vertebrate, in which HDL is the dominant plasma lipoprotein. In addition, the findings confirm the importance of fish kidney as a major site of endocytosis of macromolecules, of both exogenous and endogenous origin.