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Imaging flow cytometry (IFC) provides single‐cell imaging data at a high acquisition rate. It is increasingly used in image‐based profiling experiments consisting of hundreds of thousands of multi‐channel images of cells. Currently available software solutions for processing microscopy data can provide good results in downstream analysis, but are limited in efficiency and scalability, and often ill‐adapted to IFC data. In this work, we propose Scalable Cytometry Image Processing (SCIP), a Python software that efficiently processes images from IFC and standard microscopy datasets. We also propose a file format for efficiently storing IFC data. We showcase our contributions on two large‐scale microscopy and one IFC datasets, all of which are publicly available. Our results show that SCIP can extract the same kind of information as other tools, in a much shorter time and in a more scalable manner. [ABSTRACT FROM AUTHOR]

The purpose of this document is to provide guidance for establishing and maintaining growth and development of flow cytometry shared resource laboratories. While the best practices offered in this manuscript are not intended to be universal or exhaustive, they do outline key goals that should be prioritized to achieve operational excellence and meet the needs of the scientific community. Additionally, this document provides information on available technologies and software relevant to shared resource laboratories. This manuscript builds on the work of Barsky et al. 2016 published in Cytometry Part A and incorporates recent advancements in cytometric technology. A flow cytometer is a specialized piece of technology that require special care and consideration in its housing and operations. As with any scientific equipment, a thorough evaluation of the location, space requirements, auxiliary resources, and support is crucial for successful operation. This comprehensive resource has been written by past and present members of the International Society for Advancement of Cytometry (ISAC) Shared Resource Laboratory (SRL) Emerging Leaders Program https://isac-net.org/general/custom.asp?page=SRL-Emerging-Leaders with extensive expertise in managing flow cytometry SRLs from around the world in different settings including academia and industry. It is intended to assist in establishing a new flow cytometry SRL, re‐purposing an existing space into such a facility, or adding a flow cytometer to an individual lab in academia or industry. This resource reviews the available cytometry technologies, the operational requirements, and best practices in SRL staffing and management. [ABSTRACT FROM AUTHOR]

Analysis of imaging mass cytometry (IMC) data and other low‐resolution multiplexed tissue imaging technologies is often confounded by poor single‐cell segmentation and suboptimal approaches for data visualization and exploration. This can lead to inaccurate identification of cell phenotypes, states, or spatial relationships compared to reference data from single‐cell suspension technologies. To this end we have developed the "OPTimized Imaging Mass cytometry AnaLysis (OPTIMAL)" framework to benchmark any approaches for cell segmentation, parameter transformation, batch effect correction, data visualization/clustering, and spatial neighborhood analysis. Using a panel of 27 metal‐tagged antibodies recognizing well‐characterized phenotypic and functional markers to stain the same Formalin‐Fixed Paraffin Embedded (FFPE) human tonsil sample tissue microarray over 12 temporally distinct batches we tested several cell segmentation models, a range of different arcsinh cofactor parameter transformation values, 5 different dimensionality reduction algorithms, and 2 clustering methods. Finally, we assessed the optimal approach for performing neighborhood analysis. We found that single‐cell segmentation was improved by the use of an Ilastik‐derived probability map but that issues with poor segmentation were only really evident after clustering and cell type/state identification and not always evident when using "classical" bivariate data display techniques. The optimal arcsinh cofactor for parameter transformation was 1 as it maximized the statistical separation between negative and positive signal distributions and a simple Z‐score normalization step after arcsinh transformation eliminated batch effects. Of the five different dimensionality reduction approaches tested, PacMap gave the best data structure with FLOWSOM clustering out‐performing phenograph in terms of cell type identification. We also found that neighborhood analysis was influenced by the method used for finding neighboring cells with a "disc" pixel expansion outperforming a "bounding box" approach combined with the need for filtering objects based on size and image‐edge location. Importantly, OPTIMAL can be used to assess and integrate with any existing approach to IMC data analysis and, as it creates.FCS files from the segmentation output and allows for single‐cell exploration to be conducted using a wide variety of accessible software and algorithms familiar to conventional flow cytometrists. [ABSTRACT FROM AUTHOR]

Introduction: We studied the impact of the use of three‐dimensional multidetector computed tomography (3D‐MDCT) and fluoroscopy fusion on percutaneous left atrial appendage occlusion (LAAO) procedures in relation to procedure time, contrast volume, fluoroscopy time, and total radiation. Methods: This was a single‐center, prospective, single‐blinded, randomized control trial. Patients meeting criteria for LAAO were randomized to undergo LAAO with the WATCHMAN FLXTM device with and without 3D‐MDCT‐fluoroscopy fusion guidance using a prespecified protocol using computed tomography angiography for WATCHMAN FLXTM sizing, moderate sedation, and intracardiac echocardiography for procedural guidance. Results: Overall, 59 participants were randomly assigned to the fusion (n = 33) or no fusion (n = 26) groups. The median (interquartile range) age was 79 (75–83) years, 24 (41%) were female, and 55 (93%) were Caucasian. The median CHA2DS2 VASc and HASBLED scores were 5 (4–6) and 3 (3–4), respectively. At the time of the study, 51 (53%) patients were on a direct acting oral anticoagulant. There were no significant differences between the fusion and no fusion groups in procedure time (52.4 ± 15.4 vs. 56.8 ± 19.5 min, p =.36), mean contrast volume used (33.8 ± 12.0 vs. 29.6 ± 11.5 mls, p =.19), mean fluoroscopy time (31.3 ± 9.9 vs. 28.9 ± 8.7 min, p =.32), mean radiation dose (1177 ± 969 vs. 1091 ± 692 mGy, p =.70), and radiation dose product curve (23.9 ± 20.5 vs. 35.0 ± 49.1 Gy cm2, p =.29). There was no periprosthetic leak in the two groups in the immediate 1‐month postprocedure follow‐up periods. Conclusions: There was no significant difference with and without 3D‐MDCT‐fluoroscopy fusion in procedure time, contrast volume use, radiation dose, and radiation dose product. [ABSTRACT FROM AUTHOR]

Background: While there is recent data suggesting an advantage of computed tomography angiography (CTA) over transesophageal echocardiography (TEE) for preprocedural left atrial appendage closure (LAAC) planning, there is limited published experience for sizing strategies. Device sizing for LAAC may be challenging and noninvasive algorithms that improve this selection process are warranted. Objectives: We sought to evaluate the safety and the feasibility for the implementation of a novel CTA‐based sizing methodology for WATCHMAN™ FLX device in a series of patients undergoing LAAC using the TruPlan™ software package. Methods: A prospective analysis of 136 consecutive patients who underwent LAAC over a 12‐month period in a single, large academic hospital in the United States was conducted. CTA‐guided preprocedural planning and intracardiac echocardiography (ICE) was performed in all. Procedural success, adverse events, length of procedure, number of devices used, and length of stay were evaluated. Results: A total of 136 patients who underwent LAAC procedure with WATCHMAN™ FLX platform between October 1, 2020 until September 30, 2021 were included. The pre‐specified protocol using CTA and ICE was implemented in all patients (100%). Mean CHA2DS2VASc score was 4.4 ± 1.3 and the mean HAS‐BLED score was 3.9 ± 0.8. ICE‐guided 100% transseptal puncture success rate was 100% with 98.5% of overall procedural success rate. Preprocedural CTA sizing strategy accurately predicted the implanted size in 91.1% of patients. Ten patients (7.4%) required another sized device and 2 cases were aborted. At 45‐day follow‐up, only 1 patient (0.7%) had significant peri‐device leak (≥5 mm) on TEE. Conclusions: CTA‐based preprocedural sizing methodology for WATCHMAN™ FLX in LAAC was safe, feasible and associated with excellent procedural outcomes. Further studies are warranted to confirm if the features specific to TruPlan™ may reduce the number of deployment attempts, the number of devices utilized in the procedure, and the risk of complications. [ABSTRACT FROM AUTHOR]

Chronic lung allograft dysfunction (CLAD) remains the major barrier to long‐term survival after lung transplantation and improved insight into its underlying immunological mechanisms is critical to better understand the disease and to identify treatment targets. We systematically searched the electronic databases of PubMed and EMBASE for original research publications, published between January 2000 and April 2021, to comprehensively assess current evidence on effector immune cells in lung tissue and bronchoalveolar lavage fluid from lung transplant recipients with CLAD. Literature search revealed 1351 articles, 76 of which met the criteria for inclusion in our analysis. Our results illustrate significant complexity in both innate and adaptive immune cell responses in CLAD, along with presence of numerous immune cell products, including cytokines, chemokines and proteases associated with tissue remodelling. A clear link between neutrophils and eosinophils and CLAD incidence has been seen, in which eosinophils more specifically predisposed to restrictive allograft syndrome. The presence of cytotoxic and T‐helper cells in CLAD pathogenesis is well‐documented, although it is challenging to draw conclusions about their role in tissue processes from predominantly bronchoalveolar lavage data. In restrictive allograft syndrome, a more prominent humoral immune involvement with increased B cells, immunoglobulins and complement deposition is seen. Our evaluation of published studies over the last 20 years summarizes the complex multifactorial immunopathology of CLAD onset and progression. It highlights the phenotype of several key effector immune cells involved in CLAD pathogenesis, as well as the paucity of single cell resolution spatial studies in lung tissue from patients with CLAD. [ABSTRACT FROM AUTHOR]

Background: Mitochondrial dysfunction within neurons, particularly those of the substantia nigra, has been well characterized in Parkinson's disease and is considered to be related to the pathogenesis of this disorder. Dysfunction within this important organelle has been suggested to impair neuronal communication and survival; however, the reliance of astrocytes on mitochondria and the impact of their dysfunction on this essential cell type are less well characterized. Objective: This study aimed to uncover whether astrocytes harbor oxidative phosphorylation (OXPHOS) deficiencies in Parkinson's disease and whether these deficiencies are more likely to occur in astrocytes closely associated with neurons or those more distant from them. Methods: Postmortem human brain sections from patients with Parkinson's disease were subjected to imaging mass cytometry for individual astrocyte analysis of key OXPHOS proteins across all five complexes. Results: We show the variability in the astrocytic expression of mitochondrial proteins between individuals. In addition, we found that there is evidence of deficiencies in respiratory chain subunit expression within these important glia and changes, particularly in mitochondrial mass, associated with Parkinson's disease and that are not simply a consequence of advancing age. Conclusion: Our data show that astrocytes, like neurons, are susceptible to mitochondrial defects and that these could have an impact on their reactivity and ability to support neurons in Parkinson's disease. [ABSTRACT FROM AUTHOR]

Biosafety has always been an important aspect of daily work in any research institution, particularly for cytometry Shared Resources Laboratories (SRLs). SRLs are common‐use spaces that facilitate the sharing of knowledge, expertise, and ideas. This sharing inescapably involves contact and interaction of all those within this working environment on a daily basis. The current pandemic caused by SARS‐CoV‐2 has prompted the re‐evaluation of many policies governing the operations of SRLs. Here we identify and review the unique challenges SRLs face in maintaining biosafety standards, highlighting the potential risks associated with not only cytometry instrumentation and samples, but also the people working with them. We propose possible solutions to safety issues raised by the COVID‐19 pandemic and provide tools for facilities to adapt to evolving guidelines and future challenges. [ABSTRACT FROM AUTHOR]

Cytometry is playing a crucial role in addressing the COVID‐19 pandemic. In this commentary—written by a variety of stakeholders in the cytometry, immunology, and infectious disease communities—we review cytometry's role in the COVID‐19 response and discuss workflow issues critical to planning and executing effective research in this emerging field. We discuss sample procurement and processing, biosafety, technology options, data sharing, and the translation of research findings into clinical environments. © 2020 International Society for Advancement of Cytometry [ABSTRACT FROM AUTHOR]

We have developed and tested a novel ELISA‐like approach for bacterial detection based upon selective adhesion of targeted bacteria to microwells with prefabricated bacterial bioimprints. Bioimprints were produced from three bacterial species; Escherichia coli (Gram‐negative), Rhodococcus rhodochrous (Gram‐positive) and Sarcina aurantiaca (Gram‐negative), by using molding with curable silicone from dense layers of bacterial cells deposited on a glass substrate. We demonstrated that the surface functionalized whole cell bioimprints were able to selectively recognize and bind their own bacterial cell type. In order to detect target bacteria that are bound to the matching bioimprint, we also developed silica nanoparticles dual‐functionalized with (3‐glycidyloxypropyl)trimethoxysilane (GLYMO) coupled with 4‐hydroxyphenylboronic acid (4‐HPBA), SiO2NPs/GLYMO/4‐HPBA, which were further conjugated with horseradish peroxidase (HRP). Bacterial detection was demonstrated to work in the established ELISA‐like protocol using the colorimetric reaction of the conjugated HRP with 3,3′,5,5′‐tetramethylbenzidine (TMB). The bioimprints were used instead of capture antibodies and HRP‐coated dual functionalized silica nanoparticles instead of a secondary antibody with TMB as the enzyme‐converted reagent, producing a colored byproduct. This bacterial bioimprint‐based detection method does not rely on any antibodies, uses stable and inexpensive reagents, and could potentially find application for rapid diagnostics of bacterial pathogens in clinical samples at the point of care. [ABSTRACT FROM AUTHOR]

Payment for Ecosystem Service (PES) schemes is increasingly being used to obtain and drive environmental improvements. This paper describes two PES schemes, designed by Severn Trent Water (STW) to manage metaldehyde; Farm to Tap, and Product Substitution. Data collected over three years of running the schemes indicate that they can reduce the mean metaldehyde concentration at the Water Treatment Works by up to 61%; however, in‐catchment concentrations are more variable. Participant usage and behavioural surveys identify that the majority of farmers will use multiple methods to manage their slug problem. Farmer initiatives developed where rewards are paid for metaldehyde reductions, regardless of the process(es) used (Farm to Tap), result in 12% more metaldehyde reduction than when product substitution is used alone. Going forward, catchment management is the long‐term, economic and sustainable option. Schemes like Farm to Tap and Product Substitution are crucial to achieve water quality improvements and wider environmental benefits. [ABSTRACT FROM AUTHOR]

Objective: Health information technology has enabled efficient measurement of patient-reported outcomes (PROs). The National Institutes of Health Patient-Reported Outcomes Measurement Information System (PROMIS) is becoming more widely adopted for research and routine care, and some PROMIS instruments might be substituted for lengthier, legacy PRO instruments.Methods: Four PROMIS computer-adaptive testing (CAT) instruments (pain interference, physical function, sleep disturbance, and fatigue) and the Routine Assessment of Patient Index Data 3 (RAPID3), along with pain intensity and patient global assessment score, were administered to participants in the ArthritisPower registry. The RAPID3 was predicted using different combinations of these variables to create a new score (CAT-PROMIS RAPID3). Kappa statistics and Bland-Altman 95% limits of agreement were used to measure agreement between the observed versus predicted RAPID3.Results: A total of 6,154 eligible patients contributed 11,275 observations. The mean ± SD age was 52.7 ± 10.5 years, and 93% of patients were women. The median assessment times ranged from 29 seconds (PROMIS sleep disturbance) to 116 seconds (RAPID3). As single pairwise comparisons, the PROMIS CATs examined were modestly correlated (r approximately 0.4-0.7) to one other and RAPID3. Together with the pain intensity and patient global assessment, the PROMIS instruments explained a high fraction of total variance (R2 = 0.97) of the RAPID3 score. In the model with the highest agreement (κ = 0.93) between the observed RAPID3 and the CAT-PROMIS predicted RAPID3, Bland-Altman 95% limits of agreement showed minimal residual differences and no systematic biases.Conclusion: There was excellent agreement between the observed RAPID3 and predicted RAPID3 scores estimated using several PROMIS instruments. The Multidimensional Health Assessment Questionnaire and patient global assessment components of RAPID3 may be unnecessary if PROMIS scores are available. [ABSTRACT FROM AUTHOR]

Acute lymphoblastic leukemia (ALL) is the most common childhood cancer. While there are a number of well‐recognized prognostic biomarkers at diagnosis, the most powerful independent prognostic factor is the response of the leukemia to induction chemotherapy (Campana and Pui: Blood 129 (2017) 1913–1918). Given the potential for machine learning to improve precision medicine, we tested its capacity to monitor disease in children undergoing ALL treatment. Diagnostic and on‐treatment bone marrow samples were labeled with an ALL‐discriminating antibody combination and analyzed by imaging flow cytometry. Ignoring the fluorescent markers and using only features extracted from bright‐field and dark‐field cell images, a deep learning model was able to identify ALL cells at an accuracy of >88%. This antibody‐free, single cell method is cheap, quick, and could be adapted to a simple, laser‐free cytometer to allow automated, point‐of‐care testing to detect slow early responders. Adaptation to other types of leukemia is feasible, which would revolutionize residual disease monitoring. © 2020 The Authors. Cytometry Part A published by Wiley Periodicals, Inc. on behalf of International Society for Advancement of Cytometry. [ABSTRACT FROM AUTHOR]

Imaging flow cytometry (IFC) produces up to 12 spectrally distinct, information‐rich images of single cells at a throughput of 5,000 cells per second. Yet often, cell populations are still studied using manual gating, a technique that has several drawbacks, hence it would be advantageous to replace manual gating with an automated process. Ideally, this automated process would be based on stain‐free measurements, as the currently used staining techniques are expensive and potentially confounding. These stain‐free measurements originate from the brightfield and darkfield image channels, which capture transmitted and scattered light, respectively. To realize this automated, stain‐free approach, advanced machine learning (ML) methods are required. Previous works have successfully tested this approach on cell cycle phase classification with both a classical ML approach based on manually engineered features, and a deep learning (DL) approach. In this work, we compare both approaches extensively on the problem of white blood cell classification. Four human whole blood samples were assayed on an ImageStream‐X MK II imaging flow cytometer. Two samples were stained for the identification of eight white blood cell types, while two other sample sets were stained for the identification of resting and active eosinophils. For both data sets, four ML classifiers were evaluated on stain‐free imagery with stratified 5‐fold cross‐validation. On the white blood cell data set, the best obtained results were 0.778 and 0.703 balanced accuracy for classical ML and DL, respectively. On the eosinophil data set, this was 0.871 and 0.856 balanced accuracy. We conclude that classifying cell types based on only stain‐free images is possible with all four classifiers. Noteworthy, we also find that the DL approaches tested in this work do not outperform the approaches based on manually engineered features. © 2019 International Society for Advancement of Cytometry [ABSTRACT FROM AUTHOR]

White blood cell (WBC) differential counting is an established clinical routine to assess patient immune system status. Fluorescent markers and a flow cytometer are required for the current state‐of‐the‐art method for determining WBC differential counts. However, this process requires several sample preparation steps and may adversely disturb the cells. We present a novel label‐free approach using an imaging flow cytometer and machine learning algorithms, where live, unstained WBCs were classified. It achieved an average F1‐score of 97% and two subtypes of WBCs, B and T lymphocytes, were distinguished from each other with an average F1‐score of 78%, a task previously considered impossible for unlabeled samples. We provide an open‐source workflow to carry out the procedure. We validated the WBC analysis with unstained samples from 85 donors. The presented method enables robust and highly accurate identification of WBCs, minimizing the disturbance to the cells and leaving marker channels free to answer other biological questions. It also opens the door to employing machine learning for liquid biopsy, here, using the rich information in cell morphology for a wide range of diagnostics of primary blood. © 2019 The Authors. Cytometry Part A published by Wiley Periodicals, Inc. on behalf of International Society for Advancement of Cytometry. [ABSTRACT FROM AUTHOR]

Abstract: Variation in the phenology of avian taxa has long been studied to understand how a species reacts to environmental changes over both space and time. Penguins (Sphenicidae) serve as an important example of how biotic and abiotic factors influence certain stages of seabird phenology because of their large ranges and the extreme, dynamic conditions present in their Southern Ocean habitats. Here, we examined the phenology of gentoo (Pygoscelis papua) and chinstrap penguins (Pygoscelis antarctica) at 17 sites across the Scotia arc, including the first documented monitoring of phenology on the South Sandwich Islands, to determine which breeding phases are intrinsic, or rather vary across a species range and between years. We used a novel method to measure seabird breeding phenology and egg and chick survival: time‐lapse cameras. Contrary to the long‐standing theory that these phases are consistent between colonies, we found that latitude and season had a predominant influence on the length of the nest establishment, incubation, and guard durations. We observe a trend toward longer incubation times occurring farther south, where ambient temperatures are colder, which may indicate that exposure to cold slows embryo growth. Across species, in colonies located farther south, parents abandoned nests later when eggs were lost or chicks died and the latest record of eggs or chicks in the nest occurred earlier during the breeding period. The variation in both space and time observed in penguin phenology provides evidence that the duration of phases within the annual cycle of birds is not fundamental, or genetic, as previously understood. Additionally, the recorded phenology dates should inform field researchers on the best timing to count colonies at the peak of breeding, which is poorly understood. [ABSTRACT FROM AUTHOR]

The article presents the discussion on centers changing to cardiac computed tomography angiography (CTA) for planning left atrial appendage closure (LAAC). Topics include center adopted a cardiac magnetic resonance (CMR)‐based imaging strategy for LAAC planning; and conscious sedation guided by intracardiac echocardiography imaging and with a same‐day discharge protocol.

The vas::egfp transgenic zebrafish ( Danio rerio) could significantly enhance studies on the mechanisms by which environmental estrogens disrupt sexual differentiation, because the developing gonad can be visualized during early life via fluorescence detection. There are methodological challenges regarding dissecting out the gonads in early-life-stage fish, however, and transgene responses to estrogen exposure have not been tested. The authors exposed vas::egfp transgenic zebrafish and their wild-type siblings to the model estrogen 17α-ethinylestradiol (EE2; 0.62 ng/L and 3.33 ng/L measured concentrations) during sexual development (20−60 d posthatch) and used enhanced green fluorescent protein ( egfp) fluorescence to identify and dissect single gonads (at 40 d posthatch) to provide sufficient RNA for individual gene expression analyses, retaining the remaining gonad in the body cavity for histological analyses of sex and stage of development. Genotyping confirmed that all transgenic control fish were phenotypically egfp positive (showed green fluorescence). Interestingly, however, in a few transgenic fish exposed to EE2, no phenotypic egfp signal was seen, most notably for the 3.33 ng/L EE2 exposure. It was subsequently found that gonadal vasa expression was reduced by this concentration of EE2. Hepatic vitellogenin expression demonstrated that the vas::egfp and wild-type lines responded to estrogen with an equivalent sensitivity. The authors conclude that the vas::egfp zebrafish provides an enhanced and practical system for mechanistic studies on the effects of environmentally relevant concentrations of estrogens on gonad development. Environ Toxicol Chem 2014;33:602-605. © 2013 SETAC [ABSTRACT FROM AUTHOR]

Labeling nonquiescent cells with carboxyfluorescein succinimidyl ester (CFSE)-like dyes gives rise to a population width exceeding the threshold for resolving division peaks by flow cytometry. Width is a function of biological heterogeneity plus extrinsic and intrinsic error sources associated with the measurement process. Optimal cytometer performance minimizes extrinsic error, but reducing intrinsic error to the point of facilitating peak resolution requires careful fluorochrome selection and fluorescent cell sorting. In this study, we labeled the Jurkat and A549 cell lines with CFSE, CellTraceViolet (CTV), and eFluor 670 proliferation dye (EPD) to test if we could resolve division peaks in culture after reducing the labeled input widths by cell sorting. Reanalysis of the sorted populations to ascertain the level of reduction achieved always led to widths exceeding the gated limits due to the contribution of errors. Measuring detector-specific extrinsic error by sorting uniform fluorescent particles with similar spectral properties to the tracking dyes allowed us to determine the intrinsic error for each dye and cell type using a simple mathematical approach. We found that cell intrinsic error ultimately dictated whether we could resolve division peaks, and that as this increased, the required sort gate width to resolve any division peaks decreased to the point whereby issues with yield made A549 unsuitable for this approach. Finally, attempts to improve yields by setting two concurrent sort gates on the fluorescence distribution enriched for cells in different stages of the cell cycle that had nonequivalent proliferative properties in culture and thus should be practiced with caution. © 2013 International Society for Advancement of Cytometry [ABSTRACT FROM AUTHOR]

The composition of a dynamic mixture of similar 2,2′-bipyridine complexes of iron(II) bearing either an amide (5-benzylamido-2,2′-bipyridine and 5-(2-methoxyethane)amido-2,2′-bipyridine) or an ester (2,2′-bipyridine-5-carboxylic acid benzylester and 2,2′-bipyridine-5-carboxylic acid 2-methoxyethane ester) side chain have been evaluated by electrospray mass spectroscopy in acetonitrile. The time taken for the complexes to come to equilibrium appears to be dependent on the counteranion, with chloride causing a rapid redistribution of two preformed heteroleptic complexes (of the order of 1 hour), whereas the time it takes in the presence of tetrafluoroborate salts is in excess of 24 h. Similarly the final distribution of products is dependent on the anion present, with the presence of chloride, and to a lesser extent bromide, preferring three amide-functionalized ligands, and a slight preference for an appended benzyl over a methoxyethyl group. Furthermore, for the first time, this study shows that the distribution of a dynamic library of metal complexes monitored by ESI-MS can adapt following the introduction of a different anion, in this case tetrabutylammonium chloride to give the most favoured heteroleptic complex despite the increasing ionic strength of the solution. [ABSTRACT FROM AUTHOR]

Asymmetric cell division is an important mechanism for generating cellular diversity, however, techniques for measuring the distribution of fate-regulating molecules during mitosis have been hampered by a lack of objectivity, quantitation, and statistical robustness. Here we describe a novel imaging flow cytometric approach that is able to report a cells proliferative history and cell cycle position using dye dilution, pH3, and PI staining to then measure the spatial distribution of fluorescent signals during mitosis using CCD-derived imagery. Using Jurkat cells, resolution of the fluorescently labeled populations was comparable to traditional PMT based cytometers thus eliminating the need to sort cells with specific division histories for microscopy. Subdividing mitotic stages by morphology allowed us to determine the time spent in each cell cycle phase using mathematical modeling approaches. Furthermore high sample throughput allowed us to collect statistically relevant numbers of cells without the need to use blocking agents that artificially enrich for mitotic events. The fluorescent imagery was used to measure PKCζ protein and EEA-1+ endosome distribution during different mitotic phases in Jurkat cells. While telophase cells represented the favorable population for measuring asymmetry, asynchronously dividing cells spent approximately 43 seconds in this stage, explaining why they were present at such low frequencies. This necessitated the acquisition of large cell numbers. Interestingly we found that PKCζ was inherited asymmetrically in 2.5% of all telophasic events whereas endosome inheritance was significantly more symmetrical. Furthermore, molecular polarity at early mitotic phases was a poor indicator of asymmetry during telophase highlighting that, though rare, telophasic events represented the best candidates for asymmetry studies. In summary, this technique combines the spatial information afforded by fluorescence microscopy with the statistical wealth and objectivity of traditional flow cytometry, overcoming the key limitations of existing approaches for studying asymmetry during mitosis. © 2011 International Society for Advancement of Cytometry [ABSTRACT FROM AUTHOR]

T-cell development in the thymus and activation of mature T cells in secondary lymphoid organs requires the ability of cells to respond appropriately to environmental signals at multiple stages of their development. The process of thymocyte selection insures a functional T-cell repertoire, while activation of naive peripheral T cells induces proliferation, gain of effector function, and, ultimately, long-lived T-cell memory. The T-cell immune response is initiated upon engagement of the T-cell receptor (TCR) and coreceptor, CD4 or CD8, by cognate antigen/major histocompatibility complexes presented by antigen-presenting cells. TCR/coreceptor engagement induces the activation of biochemical signaling pathways that, in combination with signals from costimulator molecules and cytokine receptors, direct the outcome of the response. Activation of the src-family kinases p56lck (Lck) and p59fyn (Fyn) is central to the initiation of TCR signaling pathways. This review focuses on our current understanding of the mechanisms by which these two proteins orchestrate T-cell function. [ABSTRACT FROM AUTHOR]

The src-family kinases p56[suplck] (Lck) and p59[supfyn] (Fyn) are expressed in T cells and are among the first signaling molecules to be activated downstream of the T cell receptor (TCR). Evidence is emerging that although closely related, these signaling molecules have discrete functions during development, maintenance and activation of peripheral T cells. For example, during thymopoiesis Lck is uniquely able to provide all the signals required for pre-TCRβ selection, although Fyn can substitute for a subset of these. Positive selection of CD4 single-positive (SP) cells is also critically dependent on the expression of Lck but not Fyn, while differentiation of CD8 SP cells proceeds relatively efficiently in the absence of Lck. In naive peripheral T cells either Lck or Fyn can transmit TCR-mediated survival signals, and yet only Lck is able to trigger TCR-mediated expansion signals under conditions of lymphopenia. Stimulation of naive T cells by antigenic stimuli is also severly compromised in the absence of Lck, but more subtly impaired by the absence of Fyn. We discuss recent experiments addressing how these two src-kinase family members interface with downstream signaling pathways to regulate these diverse aspects of T cell behavior. [ABSTRACT FROM AUTHOR]

Investigates the roles of interleukins 5 and 13 in characterizing immune responses to respiratory sensitizing acid anhydrides. Production of interferon-gamma, interleukin-10 and mitogen inducible interleukin-4 by acid anhydride-activated lymph node cells; Induction of differential interleukin-5 and -13 secretion.

Concentrations of Zn and Cd were determined in growth rings of 13 ponderosa pines (Pinus ponderosa Dougl.) from two watersheds impacted by the discharge of heavy‐metal laden waste. Trees growing along rivers with contaminated sediments have greater concentrations of Zn and Cd than trees growing at uncontaminated reference sites. Concentrations of Zn and Cd in trees from the Coeur d'Alene region (Idaho, USA) decline progressively in younger growth rings. This indicates that reductions in mining activity and implementation of pollution control technologies have reduced Zn and Cd loading in this system. Samples from the Coeur d'Alene region contain Zn and Cd concentrations greater than those found trees from contaminated sites in the Lake Roosevelt (Washington, USA) region. Clearly, the effects of soil and other environmental conditions cannot be accounted for over the past 60 yr. However, dendrochemical analyses suggest that heavy‐metal sulfide wastes in the Coeur d'Alene region have a greater susceptibility to mobilization than siliceous slag wastes in Lake Roosevelt. Concentrations of Zn and Cd measured in a tree growing along an intervening river suggest that Zn and Cd from the Coeur d'Alene basin are diluted before reaching Lake Roosevelt. Radial translocation of Zn and Cd to the heartwood cannot be excluded as a possible influence on elemental distribution of Zn and Cd in ponderosa pine. Nonetheless, the data demonstrates that ponderosa pine can effectively integrate mobile Zn and Cd present in their environment during wood formation. [ABSTRACT FROM AUTHOR]

Mass Spectra of the methyl esters of ethane, n-propane, isopropane, n-butane, sec-butane and tert-butane sulphinic acids have been determined. In general, the parent ions are of low abundance and the most prevalent are the hydrocarbon ions which arise by α-cleavage. Other ions that serve to characterise these compounds are those from the CH4SO2 and HSO2 fragments. [ABSTRACT FROM AUTHOR]

Determines the effect of antimony impurity in lead arsenate insecticide on the antimony content of old orchard soils. Impact of the application of lead arsenate at high frequencies and rates on topsoil; Similarity of the biogeochemical behavior of antimony to arsenic (As) and phosphorus; Enhancement of the relative extractability of soil phosphorus due to the presence of high concentrations of soil As.