淋巴细胞改良培养法及其在细胞遗传学研究中的应用

摘要

Objective To establish a modified method for microculturing whole human blood for cytogenetic analysis.Methods A novel tube rack was designed to overcome the drawbacks of directly culturing the cells within centrifuge tubes. The fractions of human plasma, human serum and two commercial fetal bovine sera were analyzed with 1 5% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE).The influence of adding 0%, 5%, 10%, 1 5%,20%,25% and 30%autologous plasma to the culture on lymphocyte transformation rate and mitotic index (MI)was examined. Results The SDS-PAGE analysis showed a significant difference between commercial fetal bovine sera,and that the components of human plasma were similar to those of fetal bovine serum.The value of MI in lymphocyte was evidently increased along with addition of autologous plasma.However,this has exerted no significant effect on the transformation rate.With the addition of 10% autologous plasma,the MI value has become much higher than the conventional method.Conclusion A modified method was established by application of a novel tube inclined rack and optimization of whole blood inoculation.This method is easier and cheaper,and is suitable for application in clinical practice.%目的：建立一种外周血淋巴细胞的改良培养法，用于临床相关的细胞遗传学检测。方法设计新型培养架，解决离心管培养淋巴细胞的弊端；应用蛋白质电泳分析人血浆、人血清及商品化胎牛血清组分；考察0%、5%、10%、15%、20%、25%、30%自体血浆添加对淋巴细胞转化率及分裂指数的影响。结果十二烷基硫酸钠-聚丙烯酰胺凝胶电泳结果显示，商品化的胎牛血清质量差异显著，而人血浆与胎牛血清组分相似。自体血浆添加对淋巴细胞转化率影响不显著，但与分裂指数呈正相关。自体血浆在添加10%的情况下，即可取得优于常规培养法的实验结果。结论通过设计新型离心管斜面细胞培养架、优化全血接种量建立起的改良培养法，可以节约成本，简化工序，利于在基层推广。