A quantitative focus assay for titration of retroviruses that encode human granulocyte-macrophage colony-stimulating factor (GM-CSF).
A quantitative focus assay for titration of retroviruses that encode human granulocyte-macrophage colony-stimulating factor (GM-CSF).
复制标题
用于滴定编码人粒细胞巨噬细胞集落刺激因子 (GM-CSF) 的逆转录病毒的定量聚焦测定。
DOI：
10.1006/cyto.1998.0454
复制
发表时间：
1999
期刊：
影响因子：
--
通讯作者：
Feldman,RA
中科院分区：
文献类型：
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作者：
Feldman,RA
粒细胞-巨噬细胞集落刺激因子（GM-CSF）是一种造血生长因子，调节单核细胞-巨噬细胞和粒细胞的增殖、分化和效应功能。由于这种细胞因子能够增强抗原呈递细胞的免疫功能，编码GM-CSF的逆转录病毒已被用于将GM-CSF转染到鼠和人肿瘤细胞中，作为自体肿瘤疫苗策略的一部分。我们以前已经表明，NIH 3 T3细胞工程化表达功能性人GM-CSF受体（hGMR-NIH 3 T3），成为完全转化时，这些细胞在人GM-CSF的存在下孵育，而不是在缺乏。在这项研究中，我们使用这些hGM-CSF条件转化体设计了一种灵敏的焦点测定法，以滴定编码hGM-CSF的逆转录病毒，使用MFG-hGM-CSF/CRIP-CRIP作为我们的模型病毒。这种不含辅助因子的嗜中性逆转录病毒，经常用于将hGM-CSF转移到肿瘤细胞中，在我们的指示细胞系中具有相当大的转化性，显示出远高于105 FFU/ml的病毒滴度。本文所述的基于转化的测定法可快速测定hGM-CSF病毒的滴度，并可作为开发具有临床重要性的其他编码精氨酸的病毒的定量测定法的模型。
Granulocyte–macrophage colony-stimulating factor (GM-CSF) is a haematopoietic growth factor that regulates proliferation, differentiation, and effector functions of monocyte-macrophages and granulocytic cells. Because of the ability of this cytokine to enhance immune functions of antigen-presenting cells, retroviruses encoding GM-CSF have been used to transduce GM-CSF into murine and human tumour cells as part of autologous tumour vaccine strategies. We have previously shown that NIH 3T3 cells engineered to express functional human GM-CSF receptors (hGMR-NIH 3T3), become fully transformed when these cells are incubated in the presence but not in the absence of human GM-CSF. In this study we have used these hGM-CSF conditional transformants to devise a sensitive focus assay to titrate retroviruses encoding hGM-CSF, using MFG-hGM-CSF/Ψ-CRIP as our model virus. This helper-free amphotropic retrovirus, which has been frequently used to transduce hGM-CSF into tumour cells, was quite transforming in our indicator cell line, exhibiting virus titres well above 105FFU/ml. The transformation-based assay described here allows rapid determination of the titre of hGM-CSF-viruses, and may serve as a model for development of quantitative assays for other cytokine-encoding viruses of clinical importance.