High-throughput gene expression profiling of memory differentiation in primary human T cells.
High-throughput gene expression profiling of memory differentiation in primary human T cells.
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DOI：
10.1186/1471-2172-9-44
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发表时间：
2008-08-01
期刊：
影响因子：
3
通讯作者：
Stegmaier K
中科院分区：
文献类型：
--
作者：
Haining WN;Angelosanto J;Brosnahan K;Ross K;Hahn C;Russell K;Drury L;Norton S;Nadler L;Stegmaier K
幼稚T和B细胞分化为记忆淋巴细胞对于对病原体的免疫是必不可少的。这种细胞分化程序的治疗操作可以提高疫苗效力和记忆细胞的体外扩增。然而，鉴定诱导记忆分化的化合物的化学筛选受到以下限制：1）缺乏能够以高通量区分初始和记忆表型T细胞的启动子基因或功能测定，和2）初始T细胞的合适细胞系代表。在这里，我们描述了一种基于基因表达的筛选方法，该方法允许基于对应于这些分化状态的复杂基因签名来区分初级幼稚和记忆表型淋巴细胞。我们使用连接介导的扩增和荧光，珠为基础的检测系统，同时定量55个转录本代表幼稚和记忆表型签名在纯化的人T细胞群体。多基因面板的使用允许比任何组成单基因更好的分辨率。该方法是精确的，相关性以及与Affyphin微阵列数据，并可以很容易地扩大高通量。该方法为原代人T细胞中的高通量分化筛选提供了通用解决方案，其中没有单基因或功能测定可用。该筛选平台将允许识别指导幼稚人类淋巴细胞记忆分化的小分子、基因或可溶性因子。
The differentiation of naive T and B cells into memory lymphocytes is essential for immunity to pathogens. Therapeutic manipulation of this cellular differentiation program could improve vaccine efficacy and the in vitro expansion of memory cells. However, chemical screens to identify compounds that induce memory differentiation have been limited by 1) the lack of reporter-gene or functional assays that can distinguish naive and memory-phenotype T cells at high throughput and 2) a suitable cell-line representative of naive T cells. Here, we describe a method for gene-expression based screening that allows primary naive and memory-phenotype lymphocytes to be discriminated based on complex genes signatures corresponding to these differentiation states. We used ligation-mediated amplification and a fluorescent, bead-based detection system to quantify simultaneously 55 transcripts representing naive and memory-phenotype signatures in purified populations of human T cells. The use of a multi-gene panel allowed better resolution than any constituent single gene. The method was precise, correlated well with Affymetrix microarray data, and could be easily scaled up for high-throughput. This method provides a generic solution for high-throughput differentiation screens in primary human T cells where no single-gene or functional assay is available. This screening platform will allow the identification of small molecules, genes or soluble factors that direct memory differentiation in naive human lymphocytes.
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影响因子：
32.4
作者：
Hill, Jonathan A.;Feuerer, Markus;Benoist, Christophe
通讯作者：
Benoist, Christophe
影响因子：
64.5
作者：
Kaech, SM;Hemby, S;Ahmed, R
通讯作者：
Ahmed, R
影响因子：
15
作者：
Wu, X;Ding, S;Schultz, PG
通讯作者：
Schultz, PG
影响因子：
12.3
作者：
Peck, David;D Crawford, Emily;Ross, Kenneth N.;Stegmaier, Kimberly;Golub, Todd R.;Lamb, Justin
通讯作者：
Lamb, Justin
影响因子：
30.5
作者：
Kaech, SM;Ahmed, R
通讯作者：
Ahmed, R