Analysis of the specificity and selectivity of anti-EpCAM antibodies in breast cancer cell lines

The epithelial cell adhesion molecule (EpCAM) is a membrane glycoprotein that is expressed in most normal human epithelia and overexpressed in most carcinomas. Molecule is responsible for cell-to-cell adhesion and additionally participates in signaling, cell migration, proliferation and differentiation. Therefore, EpCAM has been the target of immunotherapy in clinical trials of several solid tumors. It appears to play an important role as a target for circulating tumor cells (CTCs) capturing. The aim of this study was to investigate and compare the specificity and selectivity of different anti-EpCAM antibodies in order to their usefulness for CTCs capturing. All experiments were performed in six different types of breast cancer cell lines (MCF-7, SkBr-3, T47D, CAMA-1, MDAMB-231, BT-20) and with use of three different anti-EpCAM antibodies (EBA-1, AUA-1, 9C4). The experiments revealed that investigated antibodies differ significantly regarding the specificity of EpCAM antigen binding. The most significant role in the circulating tumor cells capturing can play the EBA-1 and 9C4 anti-EpCAM antibodies as they revealed the most specific signal. The strength and specificity of reaction was dependent not only on the type of antibody but also on the type of breast cancer cell line. On the basis of the present outcomes it can be assumed that the best solution for obtaining the most specific results could be the use of mixture of different anti-EpCAM antibodies simultaneously. In conclusion, the proper selection of anti-EpCAM antibody is crucial especially when this antigen is considered as a marker for detection of circulating tumor cells

Integrative redescription of a common Arctic water bear Pilatobius recamieri (Richters, 1911)

Tardigrada are a group of microscopic metazoans that inhabit a variety of ecosystems throughout the world, including polar regions, where they are a constant element of microfauna with densities exceeding hundreds of individuals per gram of dry plant material. However, despite a long history of research and their ubiquity in tundra ecosystems, the majority of tardigrade species have limited and outdated diagnoses. One such example is Pilatobius recamieri, a common tardigrade that is widely distributed in the Arctic. The aim of this study is to redescribe this species using new material from the type locality and tools of integrative taxonomy, viz. by combining classical imaging and morphometry by light microscopy and scanning electron microscopy imaging with DNA sequencing of four markers with various mutation rates: three nuclear (18S rRNA, 28S rRNA, and ITS-2) and one mitochondrial (COI). The sequences of the three latter markers are also the first to be presented for the genus Pilatobius. This study therefore provides the first necessary step towards the verification of the geographic range of P. recamieri, which is currently assumed to be very broad. A detailed comparison of P. recamieri with Pilatobius secchii (Bertolani and Rebecchi, 1996) from Italy revealed no morphological or morphometric differences between the two species, thus we designate P. secchii as a nomen inquirendum until molecular data for the taxon become available. Finally, we propose to replace the term "lunula" in the superfamilies Hypsibioidea and Isohypsibioidea with the more appropriate "pseudolunula" to differentiate it from the true lunula in other parachelans

Temperature (latitude) and nutrient (seabird guano) effects on limno-terrestrial Tardigrada (Testechiniscus spitsbergensis and Pilatobius recamieri) body size

Surveys of terrestrial microinvertebrate morphometry, especially spatial patterns of body size at wider geographical scales, including the polar regions, are very scarce. In this study, we focused on Tardigrada, common limno-terrestrial microinvertebrates. Considering Bergmann’s rule, originally formulated for endothermic animals, we tested the hypothesis that body length of limno-terrestrial tardigrades augments with increasing latitude and decreasing temperature. Since some of our sampling areas adjoined seabird colonies, we also explored the effects of nutrients from seabird guano deposits. Individual body length of Testechiniscus spitsbergensis was measured in populations obtained from seven localities distributed along a latitudinal gradient extending from 45^{\circ}N (northern Italy) to 79^{\circ}N (northern Svalbard), and for Pilatobius recamieri from three localities in Svalbard (77^{\circ}N-80^{\circ}N). Considering both latitude and proximity to a seabird colony there were significant effects of locality on the body length of T. spitsbergensis; however, no clear pattern of increasing individual body size with increasing latitude could be detected. Immense differences in body size may be a signal for cryptic species diversity within this genus. No effect of latitude, or proximity to a seabird colony, on the body length of Arctic populations of P. recamieri was documented. Evidently, there is no tendency towards body size increase along the latitudinal gradient in either T. spitsbergensis or P. recamieri. Our study, and recent literature, indicates that larger body size in polar regions reported for several groups of micro-fauna may be a taxon-dependent response, and cannot be taken as a universally applicable rule for limnoterrestrial animals

Expression and cellular distribution of cyclin-dependent kinase 4 (Cdk4) and connexin 43 (Cx43) in porcine oocytes before and after in vitro maturation

It is recognised that connexin 43 (Cx43) and cyclin-dependent kinase 4 (Cdk4) are involved in the cumulus cell-oocyte communication via gap junctions and the control of cell cycle progress. However, little is known about their mRNA expression pattern and encoded proteins distribution in porcine oocytes during in vitro maturation (IVM). Cumulus-oocyte complexes (COCs) were collected from 31 puberal crossbred Landrace gilts and analysed for their Cdk4 and Cx43 mRNA expression using RQ-PCR and for the respective protein expression by confocal microscopic observations. An increased Cdk4 and Cx43 mRNA expression was found in oocytes after IVM (P < 0.001 and P < 0.05, respectively). Confocal microscopic observations revealed a significant increase of Cdk4 protein expression in the cytoplasm of oocytes during the maturation process. The localisation of Cx43 changed from zona pellucida before to cytoplasm of oocytes after IVM. It is supposed that the increased expression of Cdk4 and Cx43 mRNA in oocytes after IVM is linked with the accumulation of a large amount of templates during the process of oocyte maturation. The translocation especially of Cx43 from the zona pellucida into the cytoplasm may be associated with a decrease in gap junction activity in fully grown porcine oocytes. Both Cdk4 and Cx43 can be used as ‘checkpoints’ of oocyte maturation

The spring constant calibration of the piezoresistive cantilever based biosensor

Piezoresistive microcantilevers are widely applied to measurements of low forces, masses and viscosity [1]. After surface functionalization they might be used as a biochemical sensors being capable of the intermolecular force investigation. The problem is that such sensors change its mechanical properties in the environment they operate. Therefore there is a need for a high accuracy technique being capable of measuring of mechanical properties of functionalized cantilevers operating in the target environment. We suppose that such conditions meet the analysis of thermomechanical oscillation noise. In this paper the analysis of two types of cantilevers, that might be used in bioelectrochemical measurements, was performed. We determined the cantilever deflection and force sensitivity. The spring constant was measured by three different methods: the Cleveland methods [2] and the thermomechanical noise analysis. The obtained results indicate that analysis of thermomechanical excitation noise is the simplest and the most reliable method for spring constant calibration of piezoresistive cantilever based sensors

Biotope and biocenosis of cryoconite hole ecosystems on Ecology Glacier in the maritime Antarctic

Abstract: Despite recent great interest in glacier ecosystems in the continental Antarctic, little is known about their maritime counterparts. Our study presents descriptive data on cryoconite sediments and cryoconite holes on Ecology Glacier (King George Island) to accomplish three main objectives: (a) to identify main eukaryotic (algae, invertebrates) and prokaryotic (cyanobacteria) components of microbial communities; (b) to provide a “baseline” of community composition, organic matter and artificial contamination; and (c) identify key abiotic factors that might be important in community assembly. Cryoconite holes were sampled along an altitudinal gradient of Ecology Glacier in January, mid Austral Summer 2017. Cryoconite holes located in lower altitude were deeper than those located in the middle and the highest altitude. Seventeen species of algae and cyanobacteria with biomass of 0.79 to 5.37 µg/cm3 have been found in sediments. Dominant species were cyanobacterial Pseudanabaena frigida and Bacillariophyceae Microcostaus sp. Biomass of Bacillariophyceae was significantly higher than that of Chlorophyta and Cyanobacteria. We found three species of rotifers (two potentially new to science) and for the first time a glacier dwelling Acari (suspension feeder, Nanorchestes nivalis). Organic matter content ranged from 5.4% to 7.6%. Investigated artificial radionuclides included 137Cs, 238Pu, 239+240Pu and 241Am. 210Pb seems to be related to organic matter content. Overall, cryoconite holes on Ecology Glacier present unique habitats that serve as biodiversity hotspots of psychrophiles, source of organic matter, matrices for radioactivity tracking and model for observing changes in supraglacial ecosystems in Maritime Antarctic

Is oxygenation related to the decomposition of organic matter in cryoconite holes?

Cryoconite is a sediment occurring on glacier surfaces worldwide which reduces ice albedo and concentrates glacier surface meltwater into small reservoirs called cryoconite holes. It consists of mineral and biogenic matter, including active microorganisms. This study presents an experimental insight into the influence of sediment oxygenation on the cryoconite ability to produce and decomposition of organic matter. Samples were collected from five glaciers in the Arctic and the European mainland. Cryoconite from three glaciers was incubated in stagnant and mechanically mixed conditions to imitate inter-hole water–sediment mixing by meltwater occurring on glaciers in Northern Hemisphere, and its effect on oxygen profiles and organic matter content. Moreover, we investigated short-term changes of oxygen conditions in cryoconite from four glaciers in illuminated and dark conditions. An anaerobic zone was present or approaching zero oxygen in all illuminated cryoconite samples, varying in depth depending on the origin of cryoconite: from 1500 µm from Steindalsbreen (Scandinavian Peninsula) and Forni Glacier (The Alps) to 3100 µm from Russell Glacier and Longyearbreen (Arctic) after incubation. Organic matter content varied between glaciers from 6.11% on Longyearbreen to 16.36% on Russell Glacier. The mixed sediment from the Forni Glacier had less organic matter than stagnant, the sediment from Longyearbreen followed this trend, but the difference was not statistically significant, while the sediment from Ebenferner did not differ between groups. Our results have implications for the understanding of biogeochemical processes on glacier surfaces, the adaptation of organisms to changing physical conditions due to abrupt sediment mixing, but also on the estimation of productivity of supraglacial systems

Cryoconite – from minerals and organic matter to bioengineeredsediments on glacier's surfaces

Cryoconite is a mixture of mineral and organic material covering glacial ice, playing important roles in biogeochemical cycles and lowering the albedo of a glacier surface. Understanding the differences in structure of cryoconite across the globe can be important in recognizing past and future changes in supraglacial environments and ice-organisms-minerals interactions. Despite the worldwide distribution and over a century of studies, the basic characteristics of cryoconite, including its forms and geochemistry, remain poorly studied. The major purpose of our study is the presentation and description of morphological diversity, chemical and photoautotrophs composition, and organic matter content of cryoconite sampled from 33 polar and mountain glaciers around the globe. Observations revealed that cryoconite is represented by various morphologies including loose and granular forms. Granular cryoconite includes smooth, rounded, or irregularly shaped forms; with some having their surfaces covered by cyanobacteria filaments. The occurrence of granules increased with the organic matter content in cryoconite.Moreover, amajor driver of cryoconite colouringwas the concentration of organicmatter and its interplay with minerals. The structure of cyanobacteria and algae communities in cryoconite differs between glaciers, but representatives of cyanobacteria families Pseudanabaenaceae and Phormidiaceae, and algae families Mesotaeniaceae and Ulotrichaceaewere themost common. Themost of detected cyanobacterial taxa are known to produce polymeric substances (EPS) that may cement granules. Organic matter content in cryoconite varied between glaciers, ranging from 1% to 38%. The geochemistry of all the investigated samples reflected local sediment sources, except of highly concentrated Pb andHg in cryoconite collected fromEuropean glaciers near industrialized regions, corroborating cryoconite as element-specific collector and potential environmental indicator of anthropogenic activity. Our work supports a notion that cryoconite may bemore than just simple sediment and instead exhibits complex structure with relevance for biodiversity and the functioning of glacial ecosystem