免疫磁性细胞分选技术净化乳腺癌自体造血干细胞的实验研究

Effect of immunomagnetic selection in purging autologous peripheral blood progenitor cell of breast cancer patients

目的 评价免疫磁性细胞分选技术对乳腺癌患者的自体外周血干细胞 (APBSC)的净化效果。方法 对混有 1%MCF 7细胞系的非乳腺癌患者的APBSC以及 10份乳腺癌患者的APBSC同时进行免疫磁珠正净化、负净化和 - / +联合净化 ,比较CD34+ 细胞纯度、CD34+ 细胞回收率以及造血干细胞集落形成单位 (CFC)的富集倍数。建立免疫细胞组织化学方法 (ICC)和流式细胞术 (FCM)两种乳腺癌细胞的检测方法 ,评价三种净化方法的效果。比较细胞冻存时间对净化效果的影响。结果 10份乳腺癌患者的APBSC经正净化后 ,CD34+ 细胞纯度为 5 5 1% (2 5 9%～ 99 5 % )、回收率为 39 4 %(34 0 %～ 80 0 % ) ,乳腺癌细胞得到 2 1log(1 1log～ 2 7log)的净化。对 6例ICC检测阳性的APBSC进行 - / +联合净化后 ,CD34+ 细胞纯度为 6 4 3% (34 0 %～ 86 4 % )、回收率为 35 0 % (2 4 0 %～5 2 5 % ) ,乳腺癌细胞得到 3 0log(1 8log～ 3 9log)的清除 ,粒 /单系集落形成单位 (CFU GM)和爆式红系集落形成单位 (BFU E)分别得到 2 2倍和 3 1倍的富集。净化前全部肿瘤细胞阳性的APBSC ,经- / +联合净化后 ,ICC法检测仅有 1例表达阳性 ,FCM法检测肿瘤细胞数量明显减少。结论 免疫磁性分选APBSC是一种高效、快捷、简便的净化手段 ,- / +净?

Objective To evaluate the effect of immunomagnetic technique in purging of peripheral blood progenitor cell (PBPC) of breast cancer patients. Methods A sample of APBSC from non breast cancer patients mixed with 1% MCF 7 cell lines and 10 samples of APBSC from breast cancer patients were subjected to positive purging, negative purging, and positive/positive purging. The CD34+ cell purity, CD34+ cell recovery rate and the enrichment of colony form units were compared. The number of tumor cells before and after immunochemistry (ICC) and flow detected purging procedure cytometry (FCM). The influence of refriegeration on the effect of purging was analyzed. Results The TC depletion in 10 samples of PBPC of breast cancer patients after positive purging was 2.1(1.1～2.7) log steps, the CD34+ cell purity was 55.1 (25.9～99.5) %, and the CD34+ cell recovery was 39.4(34.0～80.0)%. After the 6 samples of ICC positive PBPC from breast cancer patients were subjected to negative selection and subsequent positive purging, the mean CD34 cell purity was 64.3 (34.0～86.4)%, the mean CD34+ cell recovery was 35.0 (24.0～52.5)%, the purging efficiency was 3.0(1.8～3.9) log, and the enrichment of colony forming units granulocyte macrophage (CFU GM) and burst forming unit erythrocyte (BFU E) were 2.2 and 3.1 respectively. TCs were detectable in all 6 cases prior to the purging procedure by ICC and FCM. After the -/+purging procedure, only one case was tumor cell positive by ICC and residual tumor cells could be detected by FCM, however the number of TCs was evidently decreased. Along with the extension of refriegeration time, the CD34 cell purity, CD34 cell recovery, and tumoe cell clearance rate decreased. Conclusion Immunomagnetic double negative/positive selectionis an effective, rapid and simple purging method.