Chemoenzymatic Labeling to Visualize Intercellular Contacts Using Lipidated SortaseA.

Methods to label intercellular contact have attracted attention because of their potential in cell biological and medical applications for the analysis of intercellular communications. In this study, a simple and versatile method for chemoenzymatic labeling of intercellularly contacting cells is demonstrated using a cell-surface anchoring reagent of a poly(ethylene glycol)(PEG)-lipid conjugate. The surface of each cell in the cell pairs of interest were decorated with sortase A (SrtA) and triglycine peptide that were lipidated with PEG-lipid. In the mixture of the two-cell populations, the triglycine-modified cells were enzymatically labeled with a fluorescent labeling reagent when in contact with SrtA-modified cells on a substrate. The selective labeling of the contacting cells was confirmed by confocal microscopy. The method is a promising tool for selective visualization of intercellularly contacting cells in cell mixtures for cell-cell communication analysis.

Active interfacial modifier: stabilization mechanism of water in silicone oil emulsions by peptide-silicone hybrid polymers.

We have developed hybrid amphiphilic polymers consisting of a silicone backbone modified with hydrocarbon chains and hydrolyzed silk peptides. These polymers are molecularly soluble neither in water nor in most of organic solvent, but are attractive with these solvents. We assume that this property enables the polymers to form "an independent third phase" between immiscible two liquid phases and stabilize the emulsion system, based on a fundamentally distinguishable mechanism from the approach by conventional surfactants. We have named these amphiphilic polymers "active interfacial modifier (AIM)" and studied physicochemical properties of AIM-stabilized water-in-silicon oil emulsions. The addition of AIM to a mixture of water and decamethylcyclopentasiloxane (D(5)) has achieved preparation of stable W/O emulsions (droplet size = ca. 1 microm) in a wide range of the three components, even under relatively gentle vortex mixing. Interestingly, the prepared W/O emulsions are found to be nearly genuine or quasi Newtonian fluid with low viscosity when water content is in the range from 0 to 36 wt % for the fixed weight ratio of AIM/D(5) = 6/4. This is a good piece of evidence that AIM forms the independent third phase, where the Newtonian shear occurs at the D(5)/AIM interface. The presence of AIM as third phase has also been confirmed by fluorescence probe method with confocal laser scanning microscopy. As such, AIM can activate interfaces by the least amount to cover interfaces as an independent third phase, and hence, this provides a new concept achieving a precise control of interfacial properties.

Water-in-oil emulsions prepared by peptide-silicone hybrid polymers as active interfacial modifier: effects of silicone oil species on dispersion stability of emulsions.

We have recently proposed a new general concept regarding amphiphilic materials that have been named as "active interfacial modifier (AIM)." In emulsion systems, an AIM is essentially insoluble in both water and organic solvents; however, it possesses moieties that are attracted to each of these immiscible liquid phases. Hence, an AIM practically stays just at the interface between the two phases and makes the resulting emulsion stable. In this study, the effects of silicone oil species on the dispersion stability of water-in-oil (W/O) emulsions in the presence of an AIM sample were evaluated in order to understand the destabilization mechanism in such emulsion systems. The AIM sample used in this study is an amphiphilic polymer consisting of a silicone backbone modified with hydrocarbon chains and hydrolyzed silk peptides. The Stokes equation predicts that the sedimentation velocity of water droplets dispersed in a continuous silicone oil phase simply depends on the expression (ρ - ρ0)/η assuming that the droplet size is constant (where ρ is the density of the dispersed water phase, ρ0 is the density of the continuous silicone oil phase, and η is the viscosity of the oil phase). The experimental results shown in this paper are consistent with the Stokes prediction: i.e., in the low-viscous genuine or quasi-Newtonian fluid region, the dispersion stability increases in the following order: dodecamethylpentasiloxane (DPS) < decamethylcyclopentasiloxane (D5) ≤ dodecamethylcyclohexasiloxane (D6). This order agrees well with the order obtained by using the expression (ρ - ρ0)/η as DPS > D5 > D6. This indicates that our emulsion system experiences destabilization through sedimentation, but hardly any coalescence occurs owing to the presence of an additional third phase consisting of the AIM that stabilizes the silicone oil/water interface in the emulsions.