背景:这项研究比较了在循环肿瘤细胞(CTC)捕获之前对血液样品进行预处理的全血稀释与密度梯度离心对通过基于大小的分离进行CTC分离的效率的影响。
方法:使用来自健康志愿者的加入SKBR3细胞的全血优化全血稀释方案的样品体积,稀释比,和多聚甲醛(PFA)浓度。来自健康志愿者的全血掺入SKBR3、A549或PC3细胞,和胃晚期患者的全血,食道,或者肝癌,用于比较最佳全血稀释方案与密度梯度离心的预处理。使用社会科学统计软件包的Studentt检验(SPSS版本17.0)进行所有统计评价。
结果:在健康志愿者的血液样本中,加标的SKBR3细胞回收率在5ml全血中最高,用2.5ml缓冲液稀释,并用0.2%PFA固定,并且当使用优化的全血稀释方案时,从5毫升全血添加的SKBR3、A549和PC3细胞回收率显著更高(87.67%±1.76%,79.50%±0.50%和71.83%±1.04%,分别)与密度梯度离心(46.83±1.76%,37.00±1.50%和41.00±1.50%,分别)。
BACKGROUND: This study compared whole blood dilution versus density gradient centrifugation for pre-processing blood samples prior to circulating tumor cell (CTC) capture on the efficiency of CTC separation by size-based isolation.
METHODS: Whole blood from a healthy volunteer spiked with SKBR3 cells was used to optimize the whole blood dilution protocol for sample volume, dilution ratio, and paraformaldehyde (PFA) concentration. Whole blood from healthy volunteers spiked with SKBR3, A549, or PC3 cells, and whole blood from patients with advanced gastric, esophageal, or liver cancer, was used to compare pre-processing by the optimal whole blood dilution protocol with density-gradient centrifugation. All statistical evaluations were performed using Student t test of the Statistical Package for Social Sciences (SPSS version 17.0).
RESULTS: In blood samples from healthy volunteers, spiked SKBR3 cell recovery rates were highest in 5 ml of whole blood, diluted with 2.5 ml buffer, and fixed with 0.2% PFA, and spiked SKBR3, A549, and PC3 cell recovery rates from 5 ml whole blood were significantly greater when using the optimized whole blood dilution protocol (87.67% ± 1.76%, 79.50% ± 0.50% and 71.83% ± 1.04%, respectively) compared to density-gradient centrifugation (46.83 ± 1.76%, 37.00 ± 1.50% and 41.00 ± 1.50%, respectively).