[Observation of double-stained African green monkey kidney COS-7 cells using total internal reflection double-channel fluorescence microscopy].

Using a Dual-View wavelength splitter, double-stained African green monkey kidney COS-7 cells, transfected with pEGFP-Myosin 15a and costained with Rhodamine-filopodia were observed based on an ICCD(intensified charge couple device) fluorescence micro-imaging systems. Total internal reflection fluorescence microscopy was used to observe the overexpression of Myosin 15a to the tips of the elongation filopodia. An approach to collecting fluorescence in two channels and avoiding spectra crosstalk was employed to observe Myosin 15a and filopodia distribution in African green monkey kidney COS-7 cells. High sensitivity TIRF technology and two channels imaging method provided a wide application in bio-medical studies.

[Nucleolar localization of human acetyltransferase-like protein and its expression in tumors].

OBJECTIVE: To confirm the nucleolar localization of telomerase-regulation associated protein-human N-acetyltransferase-like protein (hALP) and its associated functions. METHODS: Immunofluoresent staining and immunoelectron microscopy were used to detect the distribution of hALP in HeLa and Saos2 cells, and the co-localization of hALP and rDNA was analyzed by fluorescence in situ hybridization and immunofluoresence. RNAi was performed to further verify the nucleolar localization of hALP. A series of eukaryotic expression plasmids carrying various portions of hALP sequence were constructed and transiently transfected to HeLa and Saos2 cells. The expression of hALP in tumor tissues was stained by immunohistochemistry. RESULTS: hALP distributed predominantly in the nucleoli of HeLa and Saos2 cells, and colocalized with rDNA. Granular component was the precise distribution of hALP in the nucleolus under electron microscope. Nucleolar signals for hALP reduced significantly in cells transfected with hALP siRNA. The carboxy terminus of hALP including residues 549-834 was necessary for its nucleolar localization. hALP could be detected in the nucleoli of many kinds of tumor cells, including leiomyosarcoma, primitive neuroectodermal tumor, neuroblastoma, melanoma, prostatic cancer, and clear cell renal carcinoma. CONCLUSION: hALP is a nucleolar protein, and the nucleolar localization is mediated by its carboxy terminal domain, and hALP could be detected in the nucleoli of many tumor tissues, which is worthy of further investigation.