人新型冰冻血小板的制备方法与质量评价

Preparation Method and Quality Evaluation of Novel Frozen Human Platelets

目的:优化人新型冰冻血小板制备相关技术参数,制定人新型冰冻血小板的制备方案。方法:利用O型袋装富血小板血浆(PRP)制备人新型冰冻血小板,优化其制备过程中的关键技术参数(DMSO加入方式、孵育时间、离心条件等),并通过血常规检测、细胞凋亡率、血小板活化率及表面蛋白表达水平等评价冰冻血小板质量。结果:人新型冰冻血小板的制备方案中将离心去除上清的操作调整到血小板冰冻程序前,并当离心条件为800×g离心8 min时,离心操作对血小板影响最小。此外,离心前与DMSO孵育30 min的血小板在冰冻及解冻复融后的质量更优。采用本方案制备的人新型冰冻血小板经长期冰冻保存后各项指标仍保持稳定。结论:本研究制定了人新型冰冻血小板制备方案。在人新型冰冻血小板制备过程中,血小板与DMSO振荡孵育30 min后再经800×g离心8 min处理可提升冰冻血小板质量。

Objective:To optimize the technical parameters related to the preparation of novel frozen human platelets and formulate corresponding protocol for its preparation.Methods:Novel frozen human platelets were prepared with O-type bagged platelet-rich plasma(PRP),the key technical parameters(DMSO addition,incubation time,centrifugation conditions,etc.)of the preparation process were optimized,and the quality of the frozen platelets was evaluated by routine blood tests,apoptosis rate,platelet activation rate and surface protein expression level.Results:In the preparation protocol of novel frozen human platelets,the operation of centrifugation to remove supernatant was adjusted to before the procedure of platelets freezing,and the effect of centrifugation on platelets was minimal when the centrifugation condition was 800×g for 8 min.In addition,platelets incubated with DMSO for 30 min before centrifugation exhibited better quality after freezing and thawing.The indexes of novel frozen human platelets prepared with this protocol remained stable after long-term cryopreservation.Conclusion:The preparation technique of novel frozen human platelets was established and the protocol was formulated.It was also confirmed that the quality of frozen platelets could be improved by incubating platelets with DMSO for 30 min and then centrifuging them at 800×g for 8 min in the preparation of novel frozen human platelets.