Vaccine & Infectious Disease Institute, Fred Hutchinson Cancer Research Center, Seattle, WA, USA.
Retrovirology. 2010 Feb 16;7:12. doi: 10.1186/1742-4690-7-12.
SIV and HIV predominantly replicate in lymphoid tissue, but the study of virus specific CD8+ T cells in intact lymphoid tissue is difficult, as traditional in situ tetramer staining requires fresh tissue.
In this report, we demonstrate a novel technique using Qdot 655-conjugated peptide-MHC multimers to directly visualize SIV specific cells in cryopreserved tissue biopsies from chronically SIVmac239 infected Rhesus macaques. Qdot 655 multimers showed similar sensitivity and specificity to APC-conjugated tetramers by flow cytometry analysis, but yielded ten-fold higher signal intensity when imaged by fluorescence microscopy. Using this technique, we detected CD8+ T cells which recognize an immunodominant epitope (Gag CM9) in the spleen, lymph nodes, ileum and colon. In all these tissues, the Gag CM9 positive cells were mainly located in the extra follicular T cell zone. In the ileum and colon, we found Gag CM9 positive cells concentrated in Peyer's patches and solitary lymphoid follicles, a pattern of localization not previously described.
The use of Qdot multimers provide an anatomic and quantitative evaluation of SIV specific CD8+ T cell responses in SIV pathogenesis, and may prove useful to studies of SIV specific CD8+ T cell responses elicited by vaccines and other immunotherapies in the non-human primate model.
SIV 和 HIV 主要在淋巴组织中复制，但由于传统的原位四聚体染色需要新鲜组织，因此研究完整淋巴组织中的病毒特异性 CD8+T 细胞具有一定难度。
在本报告中，我们展示了一种使用 Qdot 655 缀合肽-MHC 多聚体的新技术，可直接可视化慢性 SIVmac239 感染恒河猴冷冻组织活检中的 SIV 特异性细胞。通过流式细胞术分析，Qdot 655 多聚体与 APC 缀合四聚体具有相似的灵敏度和特异性，但通过荧光显微镜成像时，其信号强度高十倍。使用该技术，我们在脾脏、淋巴结、回肠和结肠中检测到识别免疫优势表位（Gag CM9）的 CD8+T 细胞。在所有这些组织中，Gag CM9 阳性细胞主要位于滤泡外 T 细胞区。在回肠和结肠中，我们发现 Gag CM9 阳性细胞集中在派尔氏斑和孤立淋巴滤泡中，这种定位模式以前没有描述过。
使用 Qdot 多聚体可以对 SIV 发病机制中 SIV 特异性 CD8+T 细胞反应进行解剖学和定量评估，并且可能对非人类灵长类动物模型中疫苗和其他免疫疗法引发的 SIV 特异性 CD8+T 细胞反应的研究有用。
