细胞技术

Cell-based arrays offer powerful tools for genomics/proteomics and drug discovery, and are widely applicable for most cell lines. However, it is challenging to apply cell-based arrays for in vitro diagnosis due to limited amount of patient samples. Here, we utilized and demonstrated micr ...

Cell-based microarrays were first described by Ziauddin and Sabatini in 2001 as a novel method for performing high-throughput screens of gene function. They reported a technique whereby expression vectors containing the open reading frame (ORF) of human genes were printed onto glass mi ...

Analysis of the human genome sequence has identified thousands of putative genes with unknown function; therefore, a new tool allowing for rapid identification of gene functions is needed. Reverse transfection microarray technology, which turns a DNA microarray into a cell-based mi ...

Cell-based microarrays are a powerful technology platform for performing high-throughput screens of gene function. The approach entails printing expression vectors containing either genes or shRNAs onto a glass microscope slide or 384-well microtitre plate to form an array. The ...

Cell-based microarrays have been used for a wide variety of assays including gain-of-function, loss-of-function and compound screening. Many of these assays have employed fluorescent proteins as reporters. These fluorescent reporter proteins can be monitored in living cells but h ...

We have described a protocol for performing high-throughput immunofluorescence microscopy on microarrays of yeast cells. This approach employs immunostaining of spheroplasted yeast cells printed as high-density cell microarrays, followed by imaging using automated mic ...

The Golgi complex is the central organelle in the secretory membrane trafficking and its organization strongly depends upon the flow of coming and leaving material. The principles of cargo transfer to, through, and away from the Golgi complex were investigated in numerous studies. Howeve ...

Genome-wide screening procedures have developed into a useful tool for assigning new functions to known proteins or for identifying new interplayers in cell metabolism, especially under pathological conditions. Since primary cells reflect the physiological situation more c ...

Several human pathogenic viruses encode large genomes with often more than 100 genes. Viral pathogenicity is determined by carefully orchestrated co-operative activities of several different viral genes which trigger the phenotypic functions of the infected cells. Systemat ...

It is now recognized that mammalian mitochondrial DNA forms a higher structure called the nucleoid, corresponding to the nucleosome of nuclear DNA. Mitochondrial transcription factor A (TFAM), which was cloned as a transcription factor for mitochondrial DNA, is essential for the main ...

Defects in mtDNA replication are the principle cause of severe, heritable metabolic disorders classified as mitochondrial diseases. In vitro analysis of the biochemical mechanisms of mtDNA replication has proven to be a powerful tool for understanding the origins of mitochondrial ...

Tubulin, the microtubule building-block, is the target of numerous small molecule compounds that interfere with microtubule dynamics. Several of these ligands are in clinical use as antitumor drugs. There have been numerous studies on these molecules, with two main objectives: to dete ...

The knowledge of the thermodynamics and kinetics of drug-microtubule interaction is essential to understand the structure/affinity relationship of a given ligand family. When a ligand does not show an appropriate signal change (absorbance or fluorescence) upon binding, the exte ...

Vinca alkaloids play a vital role in chemotherapy protocols for a wide range of hematological and solid tumors. Studies of drug interactions with the drug target, tubulin or microtubules, have helped us to understand the cytotoxic and toxic effects. We present here in vivo and in vitro methods for s ...

Drugs that affect microtubule dynamics are among the most effective anticancer agents in routine clinical use. The standard assay for antimicrotubule agents observes the ability of a particular substance to affect in vitro microtubule assembly. We have modified these procedures so t ...

Taxol, the first microtubule stabilizer identified, is one of the most important new anticancer drugs to be brought to the clinic in the past 20 yr. The clinical success of Taxol? led to the development of a second-generation taxane, docetaxel (Taxotere™), and multiple third-generation taxane d ...

This method is a modification of the initial procedure employed to purify tubulin from mammalian brain. It consists of tissue homogenization, elimination of cell membranes, ammonium sulfate fractionation, and batch anion exchange, followed by selective precipitation with magn ...

Microtubule-binding proteins are conveniently divided into two large groups: MAPs (microtubule-associated proteins), which can stabilize, anchor, and/or nucleate microtubules, and motors, which use the energy of ATP hydrolysis for a variety of functions, including microtub ...

Tubulin heterogeneity within eukaryotic cells is generated by differential gene expression and posttranslational modification of α- and β-tubulin gene products, either as heterodimers or when polymerized into microtubules. The characterization of posttranslational ...

Cytoplasmic microtubules are noncovalent polymers of the protein tubulin. In the cells, the main function of microtubules is to provide tracks for organelle transport. Two experimental approaches based on fluorescence microscopy are commonly used to examine organization of mic ...