Differential response of human basophil activation markers: a multi-parameter flow cytometry approach

<p>Abstract</p> <p>Background</p> <p>Basophils are circulating cells involved in hypersensitivity reactions and allergy but many aspects of their activation, including the sensitivity to external triggering factors and the molecular aspects of cell responses, are still to be focused. In this context, polychromatic flow cytometry (PFC) is a proper tool to investigate basophil function, as it allows to distinguish the expression of several membrane markers upon activation in multiple experimental conditions. </p> <p>Methods</p> <p>Cell suspensions were prepared from leukocyte buffy coat of K2-EDTA anticoagulated blood specimens; about 1500-2500 cellular events for each tested sample, gated in the lymphocyte CD45dim area and then electronically purified as HLADRnon expressing/CD123bright, were identified as basophilic cells. Basophil activation with fMLP, anti-IgE and calcium ionophore A23187 was evaluated by studying up-regulation of the indicated membrane markers with a two-laser six-color PFC protocol.</p> <p>Results</p> <p>Following stimulation, CD63, CD13, CD45 and the ectoenzyme CD203c up-regulated their membrane expression, while CD69 did not; CD63 expression occurred immediately (within 60 sec) but only in a minority of basophils, even at optimal agonist doses (in 33% and 14% of basophils, following fMLP and anti-IgE stimulation respectively). CD203c up-regulation occurred in the whole basophil population, even in CD63non expressing cells. Dose-dependence curves revealed CD203c as a more sensitive marker than CD63, in response to fMLP but not in response to anti-IgE and to calcium ionophore.</p> <p>Conclusion</p> <p>Use of polychromatic flow cytometry allowed efficient basophil electronic purification and identification of different behaviors of the major activation markers. The simultaneous use of two markers of activation and careful choice of activator are essential steps for reliable assessment of human basophil functions.</p

Fatigue life analysis of solid elastomer-like polyurethanes

The material behaviour of polymeric materials under cyclic fatigue loads is complex and forms a vast field of research activities. Elastomer-like polyurethane materials form an excellent fit for cyclically loaded system components in many cases. The present work aims to quantify fatigue crack growth (FCG) in a high-performance, commercially available hydrolysis-resistant thermoplastic polyurethane (TPU). This TPU material is often used in water or oil hydraulics and applications in mining, tunneling, etc. due to its high resistance to abrasion and tear strength. For those heavy-duty applications, the critical lifetime is reached as soon as cracks reach a critical threshold in the TPU material. The first part of the current work illustrates the FCG analysis of the TPU material, i.e. crack propagation measurements on a Tear and Fatigue Analyzer (TFA, Coesfeld GmbH & Co. KG, Germany). Based on the TFA measurements, it is shown how the tearing energy and the FCG rate have a certain regularity at different strain levels and quite a different behaviour compared to standard rubber material. Secondly, a lifetime prediction of the TPU material is derived by means of advanced finite element analysis (FEA). By using Abaqus simulation software (Dassault Systèmes) with advanced material modeling concepts, simulations are performed under the identical conditions as the TFA experiments. The results are plotted in terms of total elastic strain energy density per element (ESEDEN) over FCG rate in the vicinity of the crack tip. In a third step, the lifetime prediction concept ESEDEN is cross-validated by comparing experimental results from a test bench that applies cyclic high strain rate loading to the TPU material with corresponding FEA. As demonstrated the ESEDEN data proves being a promising criterion for lifetime prediction of critical TPU components under cyclic loading conditions

Pros and Cons of Clinical Basophil Testing (BAT)

PURPOSE OF REVIEW: We review basophil testing by flow cytometry with an emphasis on advantages and disadvantages. RECENT FINDINGS: There are many tools available to assess the presence and severity of allergic diseases in patients. For 50 years, peripheral blood basophils have been used as tools to study these diseases. It is a very accessible cell that binds IgE antibody and secretes the classical mediators responsible for the symptoms of allergic reactions. In the last decade, an even more accessible methodology, using flow cytometry, has been developed to enhance the ability to use basophils for both mechanistic and clinical diagnostics. Basophil testing has been included in diagnostics for different forms of allergies as well as to monitor disease status. A variety of studies have begun to establish both precise methods and their clinical relevance for disease diagnosis, but there remain some important questions on how to take optimal advantage of the behaviours of basophils