No abstract

A method for monitoring heterogeneity in changes of plasma membrane potential (PMP) at an individual cell resolution while in suspension, utilizing a simple and low-cost wide-field illumination arrangement, is presented. The method is modeled via HEK-293 cell line in suspension, double stained with coumarin and oxonol (donor and acceptor), which were loaded into an array of nanoliter wells, each designed to preserve the individuality of the nontethered cell it holds during vigorous biomanipulation. Depolarization of PMP was induced by high K(+) solution, reducing the proximity between the membrane fluorophores and subsequently reducing the efficiency (E%) of resonance energy transfer between them. Spatial plots of E% were produced from both images of fluorescence intensity and polarization. The spatial resolution of E% plots seem to be higher, and their contrast greater, when calculated from the polarization, rather than from the intensity of the fluorescence.

In a previous study, it was observed that survivability was low when attempting to cryopreserve sperm cells in a nanoliter-sized droplet protected under soybean oil, in stark contrast to the high survival rates in milliliter-sized droplets. In this study, infrared spectroscopy was used to provide an estimate of the saturation concentration of water in soybean oil. By following the time evolution of the infrared absorption spectrum of water–oil mixtures, the saturation of water in soybean oil was found to reach equilibrium after 1 h. From the absorption spectra of neat water and neat soybean oil and the application of the Beer–Lambert law to an estimation of the absorption of a mixture from its individual components, it was estimated that the saturation concentration of water is 0.010 M. This estimate was supported by molecular modeling using the latest semiempirical methods (in particular, GFN2-xTB). While for most applications the very low solubility has little impact, the implications in those exceptions were discussed.