活体成像分析异硫氰酸荧光素标记Rituximab在荷淋巴瘤裸鼠体内的生物分布

目的 应用活体动物体内光学成像系统观察抗体药物Rituximab在荷淋巴瘤裸鼠体内的生物分布.方法 制备FITC标记的Rituximab(FITC-Rituximab),用激光扫描共聚焦显微镜和流式细胞仪体外分析FITC-Rituximab与人淋巴瘤Raji细胞的亲和力;建立裸鼠淋巴瘤皮下移植瘤模型,应用活体动物体内光学成像系统观察FITC-Rituximab在荷瘤小鼠体内的分布.结果 流式细胞仪和激光扫描共聚焦显微镜检测证明FITC-Rituximab与淋巴瘤Raji细胞有较好的亲合活性,主要结合在细胞膜表面.体内活体动物光学成像分析表明,FITC-Rituximab在1 h内即可特异性地在肿瘤部位富集,3～4 h即可进入肿瘤组织内部并达到最大浓度富集,8～10 h后在肿瘤组织仍可观察到FITCRituximab的特异性的富集,而在其他组织器官没有可见的荧光存在;双侧皮下移植瘤模型再次证明FITC-Rituximab具有对CD20抗原过表达的淋巴瘤特异性结合能力.结论 动物活体成像系统能够准确地监测FITC标记Rituximab在荷瘤裸鼠体内的动态分布情况,该技术对实时分析抗体药物在荷瘤小鼠体内的靶向效果具有参考价值和指导意义.

In vivo imaging analysis of biodistribution of FITC-labeled Rituximab in lymphoma-bearing nude mice

Objective To conduct an in vivo optical imaging analysis of the biodistribution of antibody Rituximab in lymphoma tumor-bearing nude mice. Methods Laser scanning confocal microscope and flow cytometry were employed to determine the affinity of FITC-labeled Rituximab (FITC-Rituximab)with human lymphoma Raji cells. And the in vivo optical imaging system was used to analyze the biodistribution of FITC-Rituximab in lymphoma-transplanted xenograft nude mice. Results The results of flow cytometry and laser scanning confocal microscope demonstrated that FITC-Rituximab had remarkable affinity with lymphoma Raji cells and was mainly bound at cell membrane. The results of in vivo imaging analysis suggested that FITC-Rituximab could specifically accumulated at peritumor tissue less than 1 h, then penetrated into the interior of tumor and concentrated in 3-4 h. And the specific concentration of FITCRituximab could still been observed more than 8-10 h whereas there was no apparent fluorescence at other tissues. Furthermore, the results observed from a two-flank tumor xenograft model showed that FITCRituximab possessed specific binding affinity for CD20-overexpressed lymphoma. Conclusion The in vivo optical imaging system can accurately monitor the distribution of FITC-Rituximab in tumor-bearing nude mice. And this technique has a reference value and significance for a real-time analysis of tumor-targeting capability of antibody drugs.