Steitz Julia, Tüting Thomas
Institute for Laboratory Animal Science, University Hospital of RWTH Aachen, Aachen, Germany.
Methods Mol Biol. 2013;940:317-37. doi: 10.1007/978-1-62703-110-3_24.
We describe here the use of particle-mediated gene transfer for the induction of immune responses against melanoma antigens in murine tumor models using the melanocyte differentiation antigen tyrosinase-related protein 2 (TRP2) as an antigen in a murine B16 melanoma model. We have utilized marker genes such as β-galactosidase (βgal) and EGFP, which can be readily detected, as control antigens to establish the gene delivery and to detect antigen-specific humoral and cellular immune responses. After biolistic DNA vaccination with plasmids encoding the TRP2 gene we observed the induction of TRP2-specific T-cells and antibodies associated with vitiligo-like fur depigmentation and tumor immunity against B16 melanoma cells. Here we describe the preparation of cartridges with DNA-coated gold beads and the in vivo gene transfer into skin using the Helios Gene Gun system. We also describe protocols for the measurement of humoral and cellular immune responses against the melanocyte differentiation antigen TRP2. These protocols can subsequently be adapted to other antigens.
我们在此描述在小鼠肿瘤模型中使用粒子介导的基因转移来诱导针对黑色素瘤抗原的免疫反应，在小鼠B16黑色素瘤模型中，使用黑素细胞分化抗原酪氨酸酶相关蛋白2（TRP2）作为抗原。我们利用了易于检测的标记基因，如β-半乳糖苷酶（βgal）和EGFP，作为对照抗原，以建立基因传递并检测抗原特异性体液免疫和细胞免疫反应。在用编码TRP2基因的质粒进行生物弹道DNA疫苗接种后，我们观察到诱导了TRP2特异性T细胞以及与白癜风样皮毛色素脱失和针对B16黑色素瘤细胞的肿瘤免疫相关的抗体。在此我们描述了用DNA包被的金珠制备弹夹以及使用Helios基因枪系统将基因体内转移到皮肤中的方法。我们还描述了针对黑素细胞分化抗原TRP2的体液免疫和细胞免疫反应的测量方案。这些方案随后可适用于其他抗原。
