Affinity purification of novel bispecific antibodies recognising carcinoembryonic antigen and doxorubicin

We have developed a method which combines Protein A affinity chromatography and HPLC analytical and semi-preparative hydroxyapatite affinity chromatography to purify bispecific antibodies (BsMabs) from hybrid-hybridomas secreting antibodies recognising carcinoembryonic antigen (CEA) and the chemotherapeutic drug doxorubicin (Dox). Elution of the HPLC hydroxyapatite columns with a 60–360 mM phosphate buffer gradient was found to give better separation than elution with a 60–180 mM phosphate buffer gradient. Careful monitoring of HPLC fractions by enzyme linked immunosorbent assays for anti-CEA, anti-Dox and dual anti-CEA/anti-Dox activity, and pooling of fractions on the basis of these results, enabled the purification of novel BsMabs for use in in vitro and preclinical in vivo experiments.     

Mapping the Abundance and Distribution of Adélie Penguins Using Landsat-7: First Steps towards an Integrated Multi-Sensor Pipeline for Tracking Populations at the Continental Scale

The last several years have seen an increased interest in the use of remote sensing to identify the location of penguin colonies in Antarctica, and the estimation of the abundance of breeding pairs contained therein. High-resolution (sub-meter) commercial satellite imagery (e.g., Worldview-1, Quickbird) is capable of colony detection and abundance estimation for both large and small colonies, and has already been used in a continental-scale survey of Adélie penguins. Medium-resolution Landsat imagery has been used successfully to detect the presence of breeding penguins, but has not been used previously for abundance estimation nor evaluated in terms of its minimum colony size detection threshold. We report on the first comprehensive analysis of the performance of these two methods for both detection and abundance estimation, identify the sensor-specific failure modes that can lead to both false positives and false negatives, and compare the abundance estimates of each method over multiple spatial scales. We find that errors of omission using Landsat imagery are low for colonies larger than ∼10,000 breeding pairs. Both high-resolution and Landsat imagery can be used to obtain unbiased estimates of abundance, and while Landsat-derived abundance estimates have high variance for individual breeding colonies relative to estimates derived from high-resolution imagery, this difference declines as the spatial domain of interest is increased. At the continental scale, abundance estimates using the two methods are roughly equivalent. Our comparison of these two methods represents a bridge between the more developed high-resolution imagery, which can be expensive to obtain, and the medium-resolution Landsat-7 record, which is freely available; this comparison of methodologies represents an essential step towards integration of these disparate sources of data for regional assessments of Adélie population abundance and distribution.     

In Vitro Screening for Anti-Cholinesterase and Antioxidant Activity of Methanolic Extracts of Ayurvedic Medicinal Plants Used for Cognitive Disorders

Inhibition of Acetylcholinesterase (AChE) is still considered as the main therapeutic strategy against Alzheimer’s disease (AD). Many plant derived phytochemicals have shown AChE inhibitory activity in addition to the currently approved drugs for AD. In the present study, methanolic extracts of 20 plants used in Indian Ayurvedic system of medicine for improving cognitive function were screened for acetylcholinesterase inhibitory activity by Ellman’s microplate colorimetric method. Out of 20 extracts, Emblica officinalis, Nardostachys jatamansi, Nelumbo nucifera, Punica granatum and Raulfia Serpentina showed IC₅₀ values <100 µg/ml for acetylcholinesterase inhibitory activity. Antioxidant activities of these plants were assessed by DPPH scavenging assay. Among the extracts used, antioxidant activity was highest for Terminalia chebula and Emblica officinalis with IC₅₀ values <10 µg/ml. Considering the complex multifactorial etiology of AD, these plant extracts will be safer and better candidates for the future disease modifying therapies against this devastating disease.     

Tamoxifen‐inducible podocyte‐specific iCre recombinase transgenic mouse provides a simple approach for modulation of podocytes in vivo

We report the generation and initial characterization of a mouse line expressing tamoxifen‐inducible improved Cre (iCre) recombinase (iCre‐ER^T2) under the regulation of NPHS2 (podocin) gene promoter. The resulting transgenic mouse line was named podocin‐iCreER^T2 mice. The efficiency of iCre activity was confirmed by crossing podocin‐iCreER^T2 with the ROSA26 reporter mouse. By using the floxed ROSA reporter mice, we found that tamoxifen specifically induced recombination in the kidneys. In the absence of tamoxifen, recombination was undetectable in podocin‐iCreER^T2;ROSA26 mice. However, following intraperitoneal injection of tamoxifen, selective recombination was observed in the podocytes of adult animals. We further examined the efficiency of recombination by assessing various tamoxifen exposure regimens in adult mice. These results suggest that podocin‐iCre‐ER^T2 mouse provides an excellent genetic tool to examine the function of candidate genes in podocytes in a spatially and temporally‐restricted manner. genesis 48:446–451, 2010. © 2010 Wiley‐Liss, Inc.     

A Small Genomic Region Containing Several Loci Required for Gastrulation in Drosophila

Genetic screens in Drosophila designed to search for loci involved in gastrulation have identified four regions of the genome that are required zygotically for the formation of the ventral furrow. For three of these, the genes responsible for the mutant phenotypes have been found. We now describe a genetic characterization of the fourth region, which encompasses the cytogenetic interval 24C3-25B, and the mapping of genes involved in gastrulation in this region. We have determined the precise breakpoints of several existing deficiencies and have generated new deficiencies. Our results show that the region contains at least three different loci associated with gastrulation effects. One maternal effect gene involved in ventral furrow formation maps at 24F but could not be identified. For a second maternal effect gene which is required for germ band extension, we identify a candidate gene, CG31660, which encodes a G protein coupled receptor. Finally, one gene acts zygotically in ventral furrow formation and we identify it as Traf4.     

Investigation of copper inhibition of nitrifying moving bed biofilm (MBBR) reactors during long term operations

Bioprocess and Biosystems Engineering - Copper, a prevalent heavy metal in industrial mining wastewaters, has been shown to inhibit nitrification in wastewater treatment systems. Biofilm treatment...     

Body size and composition and risk of site-specific cancers in the UK Biobank and large international consortia: A mendelian randomisation study

BackgroundEvidence for the impact of body size and composition on cancer risk is limited. This mendelian randomisation (MR) study investigates evidence supporting causal relationships of body mass index (BMI), fat mass index (FMI), fat-free mass index (FFMI), and height with cancer risk.Methods and findingsSingle nucleotide polymorphisms (SNPs) were used as instrumental variables for BMI (312 SNPs), FMI (577 SNPs), FFMI (577 SNPs), and height (293 SNPs). Associations of the genetic variants with 22 site-specific cancers and overall cancer were estimated in 367,561 individuals from the UK Biobank (UKBB) and with lung, breast, ovarian, uterine, and prostate cancer in large international consortia. In the UKBB, genetically predicted BMI was positively associated with overall cancer (odds ratio [OR] per 1 kg/m² increase 1.01, 95% confidence interval [CI] 1.00–1.02; p = 0.043); several digestive system cancers: stomach (OR 1.13, 95% CI 1.06–1.21; p < 0.001), esophagus (OR 1.10, 95% CI 1.03, 1.17; p = 0.003), liver (OR 1.13, 95% CI 1.03–1.25; p = 0.012), and pancreas (OR 1.06, 95% CI 1.01–1.12; p = 0.016); and lung cancer (OR 1.08, 95% CI 1.04–1.12; p < 0.001). For sex-specific cancers, genetically predicted elevated BMI was associated with an increased risk of uterine cancer (OR 1.10, 95% CI 1.05–1.15; p < 0.001) and with a lower risk of prostate cancer (OR 0.97, 95% CI 0.94–0.99; p = 0.009). When dividing cancers into digestive system versus non-digestive system, genetically predicted BMI was positively associated with digestive system cancers (OR 1.04, 95% CI 1.02–1.06; p < 0.001) but not with non-digestive system cancers (OR 1.01, 95% CI 0.99–1.02; p = 0.369). Genetically predicted FMI was positively associated with liver, pancreatic, and lung cancer and inversely associated with melanoma and prostate cancer. Genetically predicted FFMI was positively associated with non-Hodgkin lymphoma and melanoma. Genetically predicted height was associated with increased risk of overall cancer (OR per 1 standard deviation increase 1.09; 95% CI 1.05–1.12; p < 0.001) and multiple site-specific cancers. Similar results were observed in analyses using the weighted median and MR–Egger methods. Results based on consortium data confirmed the positive associations between BMI and lung and uterine cancer risk as well as the inverse association between BMI and prostate cancer, and, additionally, showed an inverse association between genetically predicted BMI and breast cancer. The main limitations are the assumption that genetic associations with cancer outcomes are mediated via the proposed risk factors and that estimates for some lower frequency cancer types are subject to low precision.ConclusionsOur results show that the evidence for BMI as a causal risk factor for cancer is mixed. We find that BMI has a consistent causal role in increasing risk of digestive system cancers and a role for sex-specific cancers with inconsistent directions of effect. In contrast, increased height appears to have a consistent risk-increasing effect on overall and site-specific cancers.

Mathew Vithayathil and colleagues study associations of body mass index and other measures with incidence of specific cancers.     

Recombinant monoclonal antibody yield in transgenic tobacco plants is affected by the wounding response via an ethylene dependent mechanism

Variability in recombinant IgG yield in transgenic tobacco plants has previously been observed in relation to leaf position, and is interpreted as a function of ageing and the senescence process, leading to increasing protein degradation. Here, similar findings are demonstrated in plants of different ages, expressing IgG but not IgG-HDEL, an antibody form that accumulates within the endoplasmic reticulum. Antibody yields declined following wounding in young transgenic plants expressing IgG but not in those expressing IgG-HDEL. However, in mature IgG plants, the opposite was demonstrated, with significant boosts in yield, while mature IgG-HDEL plants could not be boosted. The lack of response in IgG-HDEL plants suggests that the changes induced by wounding occur post-translationally, and the findings might be explained by wounding responses that differ in plants according to their developmental stages. Plant mechanisms involved in senescence and wounding overlap to a significant degree and compounds such as ethylene, jasmonic acid and salicylic acid are important for mediating downstream effects. Treatment of transgenic plants with ethylene also resulted in a decrease in recombinant IgG yield, which was consistent with the finding that wounded plants could induce lower IgG yields in neighbouring non-wounded plants. Treatment with 1-MCP, an ethylene antagonist, abrogated the IgG yield drop that resulted from wounding, but had no effect on the more gradual IgG yield loss associated with increasing plant age.     

Population and behavioural responses of native prey to alien predation

The introduction of invasive alien predators often has catastrophic effects on populations of naïve native prey, but in situations where prey survive the initial impact a predator may act as a strong selective agent for prey that can discriminate and avoid it. Using two common species of Australian small mammals that have persisted in the presence of an alien predator, the European red fox Vulpes vulpes, for over a century, we hypothesised that populations of both would perform better where the activity of the predator was low than where it was high and that prey individuals would avoid signs of the predator’s presence. We found no difference in prey abundance in sites with high and low fox activity, but survival of one species—the bush rat Rattus fuscipes—was almost twofold higher where fox activity was low. Juvenile, but not adult rats, avoided fox odour on traps, as did individuals of the second prey species, the brown antechinus, Antechinus stuartii. Both species also showed reduced activity at foraging trays bearing fox odour in giving-up density (GUD) experiments, although GUDs and avoidance of fox odour declined over time. Young rats avoided fox odour more strongly where fox activity was high than where it was low, but neither adult R. fuscipes nor A. stuartii responded differently to different levels of fox activity. Conservation managers often attempt to eliminate alien predators or to protect predator-naïve prey in protected reserves. Our results suggest that, if predator pressure can be reduced, otherwise susceptible prey may survive the initial impact of an alien predator, and experience selection to discriminate cues to its presence and avoid it over the longer term. Although predator reduction is often feasible, identifying the level of reduction that will conserve prey and allow selection for avoidance remains an important challenge.