Asymmetry of Cell Division in CFSE-Based Lymphocyte Proliferation Analysis

Flow cytometry-based analysis of lymphocyte division using carboxyfluorescein succinimidyl ester (CFSE) dye dilution permits acquisition of data describing cellular proliferation and differentiation. For example, CFSE histogram data enable quantitative insight into cellular turnover rates by applying mathematical models and parameter estimation techniques. Several mathematical models have been developed using different types of deterministic or stochastic approaches. However, analysis of CFSE proliferation assays is based on the premise that the label is halved in the two daughter cells. Importantly, asymmetry of protein distribution in lymphocyte division is a basic biological feature of cell division with the degree of the asymmetry depending on various factors. Here, we review the recent literature on asymmetric lymphocyte division and CFSE-based lymphocyte proliferation analysis. We suggest that division- and label-structured mathematical models describing CFSE-based cell proliferation should take into account asymmetry and time-lag in cell proliferation. Utilization of improved modeling algorithms will permit straightforward quantification of essential parameters describing the performance of activated lymphocytes

Electrokinetic characterization of extracellular vesicles with capillary electrophoresis : A new tool for their identification and quantification

This work reports on the development of the first capillary electrophoresis methodology for the elucidation of extracellular vesicles' (EVs) electrokinetic distributions. The approach is based on capillary electrophoresis coupled with laser-induced fluorescent (LIF) detection for the identification and quantification of EVs after their isolation. Sensitive detection of these nanometric entities was possible thanks to an 'inorganic-species-free' background electrolyte. This electrolyte was made up of weakly charged molecules at very high concentrations to stabilize EVs, and an intra-membrane labelling approach was used to prevent EV morphology modification. The limit of detection for EVs achieved using the developed CE-LIF method reached 8 x 10(9) EV/mL, whereas the calibration curve was acquired from 1.22 x 10(10) to 1.20 x 10(11) EV/mL. The CE-LIF approach was applied to provide the electrokinetic distributions of various EVs of animal and human origins, and visualize different EV subpopulations from our recently developed high-yield EV isolation method. (C) 2020 Elsevier B.V. All rights reserved.Peer reviewe

Quantifying CFSE label decay in flow cytometry data

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Fabrication of Novel In-Situ Remediation Tools for Unconventional Oil Contamination

The aftermath of unconventional oil (UO) accidents highlights the lack of preparedness of governments to deal with UO emergencies. Because bioremediation is considered slow process, physicochemical treatment processes are necessary in removing contaminants to constrain the spread of oil. In preliminary phase of study, bed systems for adsorption of oil compounds packed with modified dolomite were applied as pre-treatment for bioremediation systems. The high affinity of oil molecules to the active sites due to hydrophobic nature of dolomite surface, as well as low solubility of oil in water, resulted in rapid process of oil adsorption on external surface of modified dolomite. UO contaminated site contain high concentration of polyaromatic hydrocarbons (PAHs). Thus, the final phase of study focused on finding enzyme mixture for biodegradation of PAHs contaminated sites for water and soil treatment. In this regard, screening of indigenous bacteria, identification of involved enzymes, and biodegradation tests were carried out. Several combinations of the pre-selected strains were used to create most prompting consortium for enzyme production. To mimic in situ application of enzyme mixture, bioremediation of pyrene contaminated soil was carried out in soil column tests. The average values of pyrene removal after 6 weeks indicated that the enzyme cocktail can be an appropriate concentration for soil enzymatic bioremediation in the soil column system. A bioinspired device was fabricated as a sustainable remedial method. Our results showed that after 200 seconds of circulating the enzyme solution 100% of anthracene in 1.5 L of 4.6 mg/L was removed from the beaker side. In addition to the circulation of PAH degrading enzymes in hollow fiber lumens, aliphatic degrading enzymes confined in multilayer nanofibrous membrane systems play an important role in the removal of oily compounds. Based on our studies, modified polyimide aerogels were suitable to support enzyme immobilization. The degradation tests clearly showed that immobilized enzymes had biodegradation ability for model substrate in contaminated water. Our results confirmed that immobilization of cocktail enzyme mixture enhanced their storage stability, more than 45% of its residual activity at 15 ± 1 ºC for 16 days. This study could set the guideline for the enzymatic bioremediation of aromatic pollutants especially polycyclic aromatic hydrocarbons in highly contaminated soil and water body