流式细胞分选仪高效分选条件的探索

Conditions for high efficient sorting experiments by flow cytometry

目的针对如何提高流式分选试验分选得率和纯度的问题,建立高效的流式细胞分选条件,为流式分选技术应用提供参考依据。方法2019年10月至2020年6月设计以BD FACS Sorp Aria II为例,首先以微球样品作为研究对象,经检测不同分选速率、不同分选模式及不同喷嘴条件下的分选结果,分析其影响因素,并以优化的条件尝试高效分选naïve T细胞。结果70μm喷嘴,purity模式下,flow rate 1.0、3.0、5.0、7.04种速率的分选纯度均达到98%以上,但flow rate值越高,分选得率越低。不同分选模式分选时,yield模式的分选效率和得率都是最高的,分别为(99.67±0.33)%、(79.78±7.14)%,但其纯度显著低于另2种模式(均P<0.05)。3种喷嘴条件下分选后的纯度均达到98%以上,且得率差异无统计学意义(P>0.05)。结论分选原代naïve T细胞,70μm喷嘴,flow rate1.0,purity模式条件最合适。

Objective To improve the yield and purity of sorting experiments by flow cytometry,to establish efficient sorting conditions,and to provide references for the application of fluorescence-activated cell sorting.Methods The beads were used as the test samples,and the BD FACS Sorp Aria II was used as the test instrument from October,2019 to June,2020.Different sorting velocities,sorting modes,and nozzles were investigated,and the influencing factors were analyzed.The naïve T cells were sorted under the optimized conditions.Results The BD FACS Sorp Aria II gained purity of more than 98%with four different sorting velocities,flow rates of 1.0,3.0,5.0,and 7.0,by using a 70μm nozzle and the purity mode.However,the higher the flow rate,the lower the sorting yield.Sorting samples by different sorting modes,the efficiency and yield of the yield sorting mode were (99.67±0.33)% and (79.78±7.14)%, respectively, which were higher than those of theother two modes, but the purity was the lowest one by the yield sorting mode (both P<0.05). Theinstrument gained a purity of more than 98% with all the three nozzles, and the yields of the threenozzles were not statistically different (P>0.05). Conclusion When naïve T cells are sorted by BDFACS Sorp Aria II flow cytometer with a 70 μm nozzle, a flow rate of 1.0, and the purity sorting modeare preferred.