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目的探索简单、无细胞外铁产生的超低微浓度菲立磁-硫酸鱼精蛋白标记骨髓间充质干细胞(MSCs)方法。方法贴壁法培养大鼠MSCs。待3代细胞汇合至80%～90%时,更换无血清培养液,根据菲立磁和硫酸鱼精蛋白的不同浓度分为4组:A组[(7.50∶1.00)μg/ml]、B组[(10.00∶1.20)μg/ml];C组[(15.00∶1.80)μl/ml]和空白对照组,加入培养液,混匀,5%CO2孵育15min,补加血清后孵育至次日。检测细胞标记率、细胞内外铁、细胞活力和标记细胞MR信号。结果 B组可有效标记大鼠MSCs,普鲁士蓝染色阳性率100%,无细胞外铁产生,铁颗粒分布于溶酶体内。4组间台盼蓝拒染率差异无统计学意义(P>0.05);体外MR GRE T2*WI序列可检测到1×104个标记细胞。结论使用超低微浓度菲立磁10.00μg/ml与鱼精蛋白1.20μg/ml可有效标记大鼠MSCs,体外MR可检测到1×104个标记细胞。
Objective To explore a simple, extracellular iron-free ultrafine concentration of phenanthrene-protamine sulfate labeled bone marrow mesenchymal stem cells (MSCs) method. Methods Adherent MSCs were cultured in vitro. When the third generation of cells confluent to 80% ~ 90%, the serum-free medium was replaced and divided into 4 groups according to the different concentrations of phenanthrenes and protamine sulfate: A group [(7.50:1.00) μg / ml], B Group [(10.00:1.20) μg / ml]; group C [(15.00: 1.80) μl / ml] and blank control group, adding culture medium, mixing, incubating in 5% CO2 for 15min, . Cell labeling rate, intracellular and extracellular iron, cell viability and MR signal of labeled cells were detected. Results Group B could effectively label rat MSCs. The positive rate of Prussian blue staining was 100%. No extracellular iron was found and iron particles were distributed in the lysosome. There was no significant difference in the rate of trypan blue exclusion between the four groups (P> 0.05). In vitro MR GRE T2 * WI sequence could detect 1 × 104 labeled cells. Conclusions MSCs can be effectively labeled with 10.00μg / ml ultrafine concentration of phenanthroline and 1.20μg / ml of protamine, and 1 × 104 labeled cells can be detected by MR in vitro.