人胚胎干细胞不同冷冻保存方法的比较

Comparison of different cryopreservation methods of human embryonic stem cells

【目的】比较人胚胎干细胞不同冷冻保存方法的复苏率,建立高效的冷冻保存方法。【方法】利用慢速冷冻、开放式和密闭式玻璃化冷冻3种方法保存人胚胎干细胞克隆,比较这3种方法的复苏效率,并通过免疫组化染色检测复苏后细胞的OCT-4表达。【结果】开放式快速冷冻方法的克隆复苏率为(97.5±5.0)%,密闭式快速冷冻方法的克隆复苏率为(96.7±5.8)%,均极显著高于慢速冻存方法(25.7±4.1)%。解冻后的胚胎干细胞克隆仍然保持胚胎干细胞的特性。【结论】开放式和密闭式玻璃化冻存方法能够高效保存人胚胎干细胞。

【Objective】 The study compared the recovery rate of human embryonic stem（hES） cells after cryopreservation using various protocols and established an effective method for hES cells cryopreservation according to the clinic standards.【Method】 hES cells were frozen with slow-cooling protocols,vitrification protocols with open pulled straws（OPS） and vitrification protocols with closed sealed straws（CS）.The recovery rates of the three methods were compared,and Immunocytochemistry was used to observe the expression of OCT-4 on hES cells after cryopreservation.【Result】 The recovery rate was（97.5±5.0）% by vitrification with OPS and（96.7±5.8）% by vitrification with CS,which was much higher than the rate of slow-cooling,（25.7±4.1）%.Positive expression of OCT-4 in the most cultured cells was detected by immunocytochemistry.【Conclusion】 Vitrification by OPS and CS methods are reliable and effective for the cryopreservation of human embryonic stem cells.