细胞技术

Hematopoietic stem cells are at the foundation of the blood system. Their study is not only relevant to the understanding of the basic cellular mechanisms of self-renewal, lineage commitment, and differentiation, but they have also been the target of intense clinical research into the caus ...

Embryonic Stem (ES) are pluripotent cells derived from the inner cell mass of blastocysts. ES cells differentiate in vitro into all kind of cells and the development of endothelial and hematopoietic cells from mouse ES cells has been especially established. As such, the in vitro differentiat ...

The study of key mechanisms and molecules involved in the regulation of hematopoiesis in mouse models has been greatly facilitated by multi-parameter flow cytometry. Subpopulations of hematopoietic stem and progenitor cells can be identified and characterized using this techni ...

With the emergence of the concept of the leukemia stem cell, assays to study them remain pivotal in understanding (leukemic) stem cell biology. Although the in vivo NOD-SCID xenotransplantation model is still the favored model of choice in most cases, this system has some limitations as well, such ...

The use of primary human cells to model cancer initiation and progression is now within the grasp of investigators. It has been nearly a decade since the first defined genetic elements were introduced into primary human epithelial and fibroblast cells to model oncogenesis. This approach has n ...

The xenotransplantation model has been instrumental for the identification and characterization of human leukemic stem cells. In this chapter we will discuss the development of the immunodeficient model in the understanding of leukemogenesis, describe the different models of i ...

Characterization of gene expression programs and pathways important for normal and cancer stem cells has become an active area of investigation. Microarray analysis of various cell populations provides an opportunity to assess genomewide expression programs to define cellul ...

Non-random chromosomal translocations can be found in about half of acute leukaemia patients and mostly lead to either over-expression of proto-oncogenes or creation of novel fusion genes. To assess the oncogenic potential and characterize the underlying mechanisms mediated by th ...

Direct analysis of T-cells of defined specificity and phenotype, without in vitro manipulation that accompanies limiting dilution analysis or other restimulation protocols for evaluating precursor frequency, provides the most accurate representation possible of in vivo e ...

Melanoma cells are considered to be immunogenic because they express melanoma-associated antigens that are recognized by autologous T-cells. The role of T-lymphocytes in the host’s immune response to cancer in general and to melanoma in particular has been studied intensively during ...

Melanomas are most frequently infiltrated by actively proliferating T-lymphocytes (1). Some of these T-cells are cytolytic and recognize peptide antigens derived from melanoma-specific antigens (2). However, with the noteworthy exception of rare immune-mediated, sponaneo ...

To determine the activity of all human Cdc25 phosphatases, two different methods are described. For assaying phosphatase activities of recombinant Cdc25 proteins produced in Escherichia coli or insect cells, a fluorimetric assay using fluorescein diphosphate (FDP) as a substrate ...

To maintain genomic integrity against various kinds of genotoxic stress, cells have multiple checkpoints in the cell cycle. When one of the cell cycle events, such as DNA synthesis, DNA repair, and chromosomal segregation, has not been successfully completed, checkpoints will delay prog ...

Small interfering RNA (siRNA) technology has emerged as a powerful genetic tool to investigate gene function in mammalian cells. Here we use siRNA to study a mediator of DNA damage-checkpoint protein 1 (MDC1), previously known as Kiaa0170 or NFBD1, in DNA damage responses. We show that MDC1 siRNA s ...

We describe here the cloning of full-length ataxia-telangiectasia mutated (ATM) cDNA and characterization of its activity. Full-length ATM cDNA is cloned into an inducible EBV-based vector (pMEP4) and its expression analyzed in a stably transfected cell line. ATM protein induction is m ...

The mitotic checkpoint prevents cells with unaligned chromosomes from prematurely exiting mitosis by inhibiting the anaphase-promoting complex/cyclosome (APC/C) from targeting key proteins for ubiquitin-mediated proteolysis. Great efforts have been made recently to i ...

The spindle-assembly checkpoint involves signaling at kinetochores, which leads to the arrest of mitotic progression in the absence of microtubule attachment or spindle tension. Here, we detail procedures for the analysis of the spindle-assembly checkpoint in adherent mammali ...

The spindle checkpoint helps to ensure the fidelity of chromosome segregation during mitosis and meiosis. In response to sister chromatids not properly attached to the mitotic spindle, this checkpoint blocks the activity of a large ubiquitin protein ligase complex, called the anapha ...

Cell cycle checkpoints play a key role in maintaining genome stability by monitoring the order and integrity of cell division events. Checkpoints induced by DNA damage function to limit the propagation of potentially deleterious mutations. The retinoblastoma tumor suppressor (RB) ...

The completion of the human genome sequence and availability of cDNA microarray technology provide new approaches to explore global cellular regulatory mechanisms. Here we present a strategy to identify genes regulated by specific transcription factors in the human genome, and app ...