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，使用组织学、免疫组织化学（IHC）和mRNA原位杂交技术进行转基因模型研究，以及正在进行的造血基因和疾病分析，以及基因治疗方案研究，表征mRNA和蛋白质原位表达的能力。正确地确定这些类型的实验范式的功能是越来越必要的。此外，这些类型的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。一个内聚、高效和协调的CMBC将大大降低成本，并增加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，评估基因转移方案的一个关键组成部分将是转基因表达的定量和生化终点的评估。细胞和分子生物学核心不仅将协助CCEMH的大量研究人员进行转基因和基因敲除小鼠的工作，而且还将为临床前和临床基因转移研究提供必要的服务。核心将提供：1)组织的标准化处理，包埋和染色用于组织学分析，2)切片，包埋（石蜡或冷冻）和免疫组化处理。此外，抗体鉴定（Western blotting）和滴度分析将用于所用抗体的特异性和质量控制。3)mRNA原位分析将在冷冻切片上进行。对产生的每个基因探针进行正义和反义探针的制作、表征和质量控制。根据研究人员的要求，可以使用35S或非放射性探针，4)细胞自旋或组织的凋亡检测将在冷冻或石蜡切片上使用TUNEL检测。如果需要，可以使用特定基因产物的抗体和TUNEL分析进行双标记实验。总之，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，与每个研究者独立执行这些分析相比，节省了大量费用。

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.