Live Cell Correlative Imaging Probes

描述（由申请人提供）：将基于1.4 nm纳米金簇标记制备探针，用于活细胞的相关荧光和电子显微镜检查，可以在不需要对细胞膜进行物理或化学破坏的情况下进行转染。将研究三种方法：通过掺入透化化学基团如乙酰氧基甲基，或通过缀合对应于蛋白质转导结构域的肽如HIV-1达特肽以促进通过细胞膜的递送，制备组合的荧光和纳米金抗体探针的膜渗透性类似物;通过靶向融合的多组氨酸标签的抗体或次氮基三乙酸基团制备靶向绿色和其它有色荧光蛋白的膜可渗透的金簇探针;以及纳米金与FlAsH或与其相应的结合肽的缀合物，所述结合肽分别靶向表达的肽或FlAsH标记的靶。这些新探针将用于哺乳动物细胞核内基因、RNA和表达蛋白的相关定位，以及细胞核内大尺度染色质结构的研究。

DESCRIPTION (provided by applicant):Probes will be prepared, based on the 1.4 nm Nanogold cluster label, for the correlative fluorescence and electron microscopy of living cells, which may be transfected without requiring physical or chemical disruption of the cell membrane for transfection. Three approaches will be investigated: preparation of membrane permeable analogs of combined fluorescent and Nanogold antibody probes either by the incorporation of permeabilizing chemical groups such as acetoxymethyl, or by the conjugation of peptides corresponding to protein transduction domains such as the HIV-1 Tat peptide to facilitate delivery through cell membranes; preparation of membrane permeable gold cluster probes targeted to green and other colored fluorescent proteins by means of antibodies or nitrilotriacetic acid groups which target fused polyhistidine tags; and conjugates of Nanogold with FlAsH or with its corresponding binding peptide targeted respectively to the expressed peptide or to FlAsH-labeled targets. The new probes will be evaluated in the correlative localization of genes, RNA and expressed proteins in the mammalian cell nucleus, and to study the large-scale chromatin structure in the nucleus.