CORE--CELL AND MOLECULAR BIOLOGY

细胞和分子生物学核心(CMBC)将为CCEMH成员提供服务，在他们的转基因模型研究中，以及在造血基因和疾病的分析中，以及在基因治疗方案的研究中，利用组织学、免疫组织化学(IHC)和mRNA原位杂交，原位表征mRNA和蛋白质表达的能力变得越来越必要，以正确确定这些类型的实验范例中的功能。此外，这类分析的数量预计将迅速增加，超过独立调查人员以具有成本效益或及时的方式进行这些分析的能力。一个有凝聚力、高效和协调的CMBC将大大降低成本，并提高为CCEMH成员执行这些分析的可靠性。此外，随着未来基因转移效率的提高，基因转移协议评估中的一个关键组成部分将是转基因表达的量化和生化终点的评估。细胞和分子生物学核心不仅将协助CCEMH的大量研究人员处理转基因和基因敲除小鼠，还将为临床前和临床基因转移研究提供必要的服务。该核心将提供：1)用于组织学分析的标准化处理、包埋和染色；2)切片、包埋(石蜡或冰冻)和IHC处理。此外，还将进行抗体鉴定(Western Blotting)和滴度分析，以确定所用抗体的特异性和质量控制。3)冰冻切片上将进行mRNA原位分析。将生产正义和反义探针，并对产生的每个基因探针进行表征和质量控制。根据研究人员的要求，将使用35S或非放射性探针，以及4)对细胞胞浆或组织进行细胞凋亡分析将在冰冻或石蜡切片上使用TUNEL分析。如果需要，可以使用针对特定基因产物的抗体以及TUNEL分析来进行双重标记实验。总而言之，该核心将提供准确、高效和具有成本效益的体内基因表达分析，与由每个个体研究人员独立执行这些分析相比，可以节省大量成本。

The Cellular and Molecular Biology Core (CMBC) will provide services to the CCEMH members using histology, immunohistochemistry (IHC) and mRNA in situ hybridization in their studies of transgenic models, as well as in the lines are being produced in the analysis of hematopoietic genes and diseases, as well as in the studies of gene therapy protocols, the ability to characterize the expression in situ, both for mRNA and protein, is becoming and more a necessity to correctly ascertain the function in these types of experimental paradigms. Furthermore, the number of these types of analyses is projected to increase rapidly, outstripping the ability of independent investigators to perform these analyses in a cost-effective or timely manner. A cohesive, efficient, and coordinated CMBC, will greatly decrease the cost and increase the reliability for these analyses to be performed for members of the CCEMH. Furthermore, as the efficiency of gene transfer improves in the future, a critical component in the evaluation of gene transfer protocols will be quantitation of transgene expression and assessment of biochemical endpoints. The Cell and Molecular Biology Core will not only assist a large number of investigators of the CCEMH working with transgenic and knock-out mice, but will also provide essential services to pre-clinical and clinical gene transfer research. The core will provide: 1) standardized processing, embedding and staining of tissues for histological analysis, 2) sectioning, embedding (paraffin or frozen) and IHC processing. Also, antibody characterization (Western blotting) and titer analysis will be performed for specificity and quality control of the antibodies used, 3) mRNA in situ analysis will be performed on frozen sections. Sense and anti-sense probes will be produced, characterized and quality controlled for each gene probe generated. Either 35S or non-radioactive probes will be used as per the investigators request, and 4) apoptosis assays on cell cytospins or tissues will be performed using the TUNEL assay on frozen or paraffin sections. Double labeling experiments can be performed using an antibody for specific gene product along with the TUNEL assay if required. Altogether, the core will provide accurate, efficient and cost-effective analysis of gene expression in vivo, with substantial savings compared to having each individuals investigator performing these analyses independently.