Hong Enping, Dobrovolskaia Marina A.
Nanotechnology Characterization Lab, Cancer Research Technology Program, Frederick National Laboratory for Cancer Research sponsored by the National Cancer Institute, Frederick, MD 21702
The stimulation of antigen-specific T-cell responses is critical for therapeutic cancer vaccines. Extensive efforts have been made to develop nanoparticles as carriers of therapeutic cancer vaccines to antigen presenting cells (APCs) of the immune system. The purpose of this protocol is to use nanoparticles to deliver vaccines to APCs and measure the antigen-specific T-cell responses that are instigated. For the purposes of this assay, the model antigen ovalbumin (OVA) is used in the context of murine cells. The protocol requires the synthesis of nanoparticles that deliver SIINFEKL (OVA), the immunodominant class I peptide derived from OVA, either in its peptide form or as part of a larger molecule (such as the whole OVA protein). CD8 T-cells that recognize SIINFEKL are purified from the transgenic OT-I mouse model [1], and activated using nanoparticle-treated APCs. The resulting T-cell proliferation, as well as secretion of the cytokines interferon gamma (IFN-γ) and interleukin-2 (IL-2), are used to measure antigen-specific T-cell activation. The results of this protocol may be used to infer the ability of the nanoparticle system to deliver other similar antigens.
抗原特异性T细胞反应的刺激对于治疗性癌症疫苗至关重要。人们已做出大量努力来开发纳米颗粒，将其作为治疗性癌症疫苗的载体，输送至免疫系统的抗原呈递细胞（APC）。本方案的目的是利用纳米颗粒将疫苗递送至APC，并测量由此引发的抗原特异性T细胞反应。为了该检测的目的，在小鼠细胞环境中使用模型抗原卵清蛋白（OVA）。该方案要求合成能递送SIINFEKL（OVA）的纳米颗粒，SIINFEKL是源自OVA的免疫显性I类肽，其形式可以是肽，也可以是更大分子（如完整OVA蛋白）的一部分。识别SIINFEKL的CD8 T细胞从转基因OT-I小鼠模型中纯化出来[1]，并使用经纳米颗粒处理的APC进行激活。由此产生的T细胞增殖以及细胞因子γ干扰素（IFN-γ）和白细胞介素-2（IL-2）的分泌，用于测量抗原特异性T细胞激活。本方案的结果可用于推断纳米颗粒系统递送其他类似抗原的能力。
