Preparation of homogeneous endoderm cells and culture is a prerequisite to understanding the cellular and molecular mechanism of endosymbiosis in the cnidarian-dinoflagellate association. During the cell isolation from the stony coral Euphyllia glabrescens, various amounts of symbiotic endoderm cells were found to release their symbionts (Symbiodinium spp., or zooxanthellae in generic usage) into the culture. Due to the bulky occupation by zooxanthellae inside the endoderm cell, the symbiotic endoderm cells, or zooxanthellae in hospite, are difficult to be distinguished from released zooxanthellae by microscopic examination. We now report a method for this identification using a fluorescent analogue of sphingomyelin, N-[5-(5,7-dimethyl boron dipyrromethene difluoride)-1-pentanoyl]-D-erythro-sphingosylphosphorylcholine (C(5)-DMB-SM). Incubation of symbiotic endoderm cells with C(5)-DMB-SM-defatted bovine serum albumin (DF-BSA) complex results in bright fluorescent membrane staining. Nevertheless, the membrane staining of free-living or released zooxanthellae by this complex is significantly decreased or even diminished. This method has provided a fast and reliable assay to identify symbiotic endoderm cells and will greatly accelerate the progress of endosymbiosis research.

Electrorotation (ER) has become a very powerful diagnostic technique for the measurement of dielectric properties of cells. However, only a few papers have investigated the electric-induced rotation of particles in a stationary alternating (AC) electric field instead of a rotating electric field. In this study, a microchip composed of a top-grounded electrode, flow chamber and bottom chess-type electrode arrays was used to construct a stationary non-uniform AC electric field for the manipulation of cells by dielectrophoretic force. We focused on the effects of metal and dielectric nanoparticles uptaken by cells under ER, by using human promyelocytic leukemia cells (HL-60), 13 nm Au and 19 nm SiO(2) nanoparticles. As revealed by the experimental results, both the percentage of cells in rotation and the range of rotational (ROT) frequency for the uptake of Au nanoparticle cells were higher and wider than in the case of SiO(2) nanoparticles. In addition, the rotation of lone cells and pearl-chain cells under non-uniform and uniform electric field were quantitatively investigated, respectively. The membrane capacitance and membrane conductance of HL-60 cells can be extracted from the ROT spectra as 10.18+/-1.92 mF/m(2) and 1500+/-321 S/m(2), respectively. In general, the ER of cells in a stationary AC electric field can be attributed to the highly non-uniform electric field and non-uniform dispersion of nanoparticles within cells; therefore, the electrical properties of uptaken nanoparticles and the aggregation phenomenon have significant influences on the resulting electrical torque.

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