定量检测残留白细胞方法的建立及在辐照血小板中的应用

Set up A Method of Quantifying Residual White Blood Cells and Application in Irradiation Platelets

目的建立定量检测单采血小板中残留白细胞的流式细胞术方法,探讨经血液辐照仪辐照前后血小板中白细胞的含量变化。方法应用绝对计数管内的定量微球(Beads)调整流式细胞仪参数,确定流式细胞仪检测条件。血小板样品经过RNA酶、固定剂、渗透剂作用后,再经碘化丙啶标记,通过流式细胞仪计数经标记的白细胞,建立检测残留白细胞含量的方法。采用血细胞分离机采集单采血小板16人份,通过血液辐照仪25Gy照射310s,应用建立的流式细胞术方法跟踪检测单采血小板辐照前及辐照后0、1、3、5d的白细胞含量,比较白细胞含量的变化。结果单采血小板中残留白细胞含量范围为1.85×106～1.3×108/袋。单采血小板中残留白细胞计数于辐照前与辐照后0d相比没有变化,与辐照后1d相比残留白细胞数量的减少有显著性(P<0.05);辐照后1d白细胞计数与辐照后3d和5d相比没有改变。结论流式细胞术方法具有较好的准确性、客观性和重复性,可以成为评估血小板质量的监控手段。辐照不立即改变血小板中残留白细胞的计数,辐照后1d残留白细胞的数量减少。

Objective To establish a method of quantifying residual white blood cells （rWBCs） in apheresis platelets by flow cytometry （FCM）, and to discuss the change of rWBCs in irradiation platelets. Methods The beads in absolute enumerative tube was used to adjust the parameter of cytometry and determine the instrument setup. After treated by RNase, fixation and permeability reagent,the rWBCs of platelet samples were stained by the PI, and detected by flow cytometry. Subsequently 16 packs apheresis platelets collected with the MCS＋ cell separator were irradiated with 25 Gy for 310 second, and quantified rWBCs using FCM method established by us in 0,1,3,5 day（s） of storage after irradiated. Results The range of rWBCs in apheresis platelets collected with MCS＋ was 1.85 ）〈 10^6- 1.3 ）〈 10^8/pack. There was no difference about the rWBC counts of platelet samples between un-irradiated ancl storage 0 day after irradiated, whereas the rWBC counts of storage 1 day significantly decreased （P〈0. 05）, and the counts of storage 1 day and 3 or 5 days after being irradiated didn＇t change. Conclusion The FCM method is highly accurate,objective and repetitive,and it would become a means to evaluate the quality of platelets. The rWBC counts don＇t alter at once by irradiation, but the counts will decrease in storage 1 day after irradiation.