LACZ INDUCIBLE, ANTIGEN MHC-SPECIFIC T-CELL HYBRIDS
LACZ INDUCIBLE, ANTIGEN MHC-SPECIFIC T-CELL HYBRIDS
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DOI：
10.1093/intimm/6.3.369
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发表时间：
1994-03-01
影响因子：
4.4
通讯作者：
SHASTRI, N
中科院分区：
文献类型：
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作者：
SANDERSON, S;SHASTRI, N
IL-2基因的活化T细胞核因子（NFAT）增强子元件可以调节大肠杆菌lacZ报告基因在活化T细胞中的表达。基于这一观察，我们表明这种诱导型NFAT-lacZ构建体可用于测量单个T细胞中TCR介导的配体特异性活化。在这里，我们描述了通过用NFAT-lacZ构建体转染α-β-BW 5147细胞产生两种新的融合伴侣BWZ. 36和BWZ. 36 CD 8 α来获得lacZ诱导型T细胞杂合体的一般方法。使用这些融合伙伴和免疫小鼠的正常T细胞，我们获得了T细胞杂交体，其中lacZ活性特异性诱导抗原/MHC II类或MHC I类复合物。我们表明，测量配体诱导的T细胞活化的非放射性lacZ测定是更简单，更快，更符合成本效益的相对于传统的IL-2测定。最重要的是，通过lacZ测定检测单个T细胞活化的独特能力允许检测罕见的抗原呈递细胞，从而为开发用于鉴定未知T细胞抗原的表达克隆策略提供基础。
The nuclear factor of activated T cells (NFAT) enhancer element of the IL-2 gene can regulate expression of the Escherichia coli lacZ reporter gene in activated T cells. Based upon this observation, we showed that this inducible NFAT - lacZ construct could be used to measure TCR mediated, ligand-specific activation in single T cells. Here we describe a general approach to obtaining lacZ inducible, T cell hybrids by generating two new fusion partners BWZ.36 and BWZ.36 CD8alpha derived by transfecting alpha-beta-BW5147 cells with the NFAT - lacZ construct. Using these fusion partners and normal T cells from immunized mice, we obtained T cell hybrids in which lacZ activity is specifically induced in response to antigen/MHC class II or MHC class I complexes. We show that measuring ligand induced T cell activation by the non-radioactive lacZ assay is simpler, faster, and more cost-effective relative to conventional IL-2 assays. Most importantly, the unique ability to detect activation of single T cells by the lacZ assays permits detection of rare antigen presenting cells and thus provides the basis for developing expression cloning strategies for identifying unknown T cell antigens.