Intracellular signal transduction relies on spatial and temporal signal transmitter dynamics. To clarify the correlations of these transmitter molecules, multicolor-imaging has been widely used. However, in the case of applying multiple indicators in a cell, spectral overlap of the indicators prevents accurate quantitative analysis. Moreover, the invasive (toxic) effect, the localization, the metabolism, as well as photobleaching of these indicators complicate the situation. Here, we show that single-molecular multifluorescent probes can overcome these problems. While intracellular calcium plays a critical role as a signal transmitter and magnesium acts as a cofactor in many situations, the correlations between the two cations are now the main issue. We designed and synthesized a Ca2+-Mg2+ responsive multifluorescent probe, KCM-1. KCM-1 shows a spectral blue shift upon complexation to Ca2+ and a red shift to the presence of Mg2+. With data analyzed at different excitation wavelengths, the concentrations of Ca2+ and Mg2+ are simultaneously quantified. Furthermore, by using the AM-ester method, intracellular Ca2+ and Mg2+ concentrations are simultaneously imaged. Such a type of intracellular multiple analyte imaging by a single-molecular multifluorescent probe is successfully demonstrated for the first time.

Synopsis :Over the last decade, regulations regarding end-of-life vehicle (ELV) recycling have been introduced in many countries, leading to an increased recycling rate of ELVs. However, steel scrap from ELVs is recycled as a secondary iron source in the steelmaking process with rare consideration of alloying elements loss. Consequently, dissipation to slag and contamination in metals with alloying elements during the steel recycling process are likely to occur, but very few open data about destination of alloying elements which is available for stakeholders of ELV recycling. The purpose of this study is to obtain information about the amounts of alloying elements associated with ELV scrap and its allocation ratio between metal and slag phases in an electric arc furnace (EAF) steel making process. Our results indicate that, over half a kilogram of chromium and manganese are contained in a unit of ELV. 10% of chromium and 20-40% of manganese are transferred to slag during the dissolution process, and more alloying elements are expected to be lost depend on the condition of oxidation pressure.