Neelapu S S, Baskar S, Kwak L W
ETI Branch, NCI at Frederick, Md 21702-1201, USA.
J Cancer Res Clin Oncol. 2001 Oct;127 Suppl 2:R14-9. doi: 10.1007/BF01470994.
Keyhole limpet hemocyanin (KLH) has been used as an immune potentiator to enhance antigen-specific responses against haptens and weak antigens including self-antigens. In the present study, we describe the optimization of the intracellular cytokine response to KLH in peripheral blood mononuclear cells (PBMC) of lymphoma and myeloma patients that were vaccinated with tumor-specific immunoglobulin (Id) conjugated to KLH. Addition of anti-CD28 antibody significantly enhanced cytokine-producing CD4-T cells. While fresh PBMC showed maximal cytokine response 14 h after antigen stimulation, the frozen PBMC showed maximal cytokine responses by 24 h. Supplementation of the culture medium with fetal bovine serum gave a better signal-to-noise ratio than human AB serum in the intracellular detection of cytokines. The intracellular cytokine responses correlated with the cytokine measurements by enzyme-linked immunosorbent assay (ELISA). Together these results indicate that the intracellular cytokine assay is very helpful to measure antigen-specific immune responses, and in subsequent studies, we have utilized this sensitive technique to detect immune responses against tumor antigens such as idiotype.
匙孔血蓝蛋白（KLH）已被用作免疫增强剂，以增强针对半抗原和包括自身抗原在内的弱抗原的抗原特异性反应。在本研究中，我们描述了淋巴瘤和骨髓瘤患者外周血单核细胞（PBMC）中对与KLH偶联的肿瘤特异性免疫球蛋白（Id）进行疫苗接种后，细胞内细胞因子对KLH反应的优化。添加抗CD28抗体可显著增强产生细胞因子的CD4-T细胞。新鲜PBMC在抗原刺激后14小时显示出最大细胞因子反应，而冷冻PBMC在24小时显示出最大细胞因子反应。在细胞内细胞因子检测中，用胎牛血清补充培养基比用人AB血清具有更好的信噪比。细胞内细胞因子反应与酶联免疫吸附测定（ELISA）的细胞因子测量结果相关。这些结果共同表明，细胞内细胞因子测定对于测量抗原特异性免疫反应非常有帮助，在随后的研究中，我们利用这种灵敏技术检测针对肿瘤抗原（如独特型）的免疫反应。
