Gerner Wilhelm, Stadler Maria, Hammer Sabine E, Klein Daniela, Saalmüller Armin
Institute of Immunology, Department of Pathobiology, University of Veterinary Medicine Vienna, Veterinärplatz 1, 1210 Vienna, Austria.
Vet Immunol Immunopathol. 2010 Nov 15;138(1-2):154-8. doi: 10.1016/j.vetimm.2010.07.009. Epub 2010 Jul 16.
The transcription factor forkhead-box p3 (Foxp3) has been designated as a master regulator for the function of regulatory T cells (Treg). Therefore, the identification of Foxp3 expression in T cells is indispensable for the study of Treg. However, studies on Foxp3 expression in bovine lymphocytes are still sparse, probably due to a lack of Foxp3-specific antibodies with reliable performance in flow cytometry. Our group recently demonstrated that a monoclonal antibody (FJK-16s) developed against murine Foxp3 also binds to porcine Foxp3 and performs well in flow cytometry. A protein sequence alignment of the binding region of the FJK-16s antibody revealed, that within this region the sequences of porcine and bovine Foxp3 are identical. Therefore, we tested this antibody for its suitability in flow cytometry with bovine peripheral blood mononuclear cells (PBMC). By using nonspecific isotype-matched antibodies and competition labeling with non-fluorescent FJK-16s antibodies as negative controls, we readily observed a specific staining of a small subpopulation of CD25(high) lymphocytes within PBMC. Co-staining with monoclonal antibodies against CD3, CD4, CD8β and TCR-γδ revealed that all Foxp3+ cells co-expressed CD3, and were in their vast majority CD4+. However, minor populations of Foxp3+CD8β+ and Foxp3+TCR-γδ+ lymphocytes could also be identified. In summary, our data demonstrate that the FJK-16s antibody is a valuable tool to promote the study of Foxp3+ T cells and their biological relevance in cattle.
转录因子叉头框蛋白p3（Foxp3）已被确定为调节性T细胞（Treg）功能的主要调节因子。因此，鉴定T细胞中Foxp3的表达对于Treg的研究必不可少。然而，关于牛淋巴细胞中Foxp3表达的研究仍然很少，这可能是由于缺乏在流式细胞术中具有可靠性能的Foxp3特异性抗体。我们小组最近证明，一种针对小鼠Foxp3开发的单克隆抗体（FJK-16s）也能与猪Foxp3结合，并在流式细胞术中表现良好。FJK-16s抗体结合区域的蛋白质序列比对显示，在该区域内猪和牛Foxp3的序列是相同的。因此，我们测试了该抗体在牛外周血单个核细胞（PBMC）流式细胞术中的适用性。通过使用非特异性同型匹配抗体和用非荧光FJK-16s抗体进行竞争标记作为阴性对照，我们很容易观察到PBMC中一小部分CD25（高）淋巴细胞的特异性染色。与抗CD3、CD4、CD8β和TCR-γδ单克隆抗体共染色显示，所有Foxp3+细胞均共表达CD3，且绝大多数为CD4+。然而，也可以鉴定出少量的Foxp3+CD8β+和Foxp3+TCR-γδ+淋巴细胞。总之，我们的数据表明，FJK-16s抗体是促进牛Foxp3+T细胞及其生物学相关性研究的有价值工具。
