平移计数法在微团培养技术中的应用

New Mensuration Methods in Micromass Rat Embryo Cell Cultures

背景与目的探索一种经济、快速而又准确的计数方法,以应用于微团培养技术中集落的评价。材料与方法采用5d连续培养妊娠13d的大鼠胚胎中脑及肢芽细胞后,以倒置显微镜视场直径为间距将微团划分为若干等间距平行区间,沿一定方向移动视场并依次计数各区间内集落数,通过摄像分析和批内重复实验评价方法的准确度及灵敏度。结果各组数据经Grubbs检验法检验均为正常数据,中脑细胞集落数和肢芽细胞集落数的相对误差分别为0.776%、0.213%;批内标准差分别为0.9339、2.8839;批内变异系数分别为1.24%、2.05%,均小于WHO提出的OCV(3%)标准。结论平移记数法无需特殊设备,不仅具有较好的准确度和极高的灵敏度,而且适用于即时分析,从而缩短实验周期,可以作为微团培养技术中集落计数的常规方法。

BACKGROUND ＆ AIM： To explore an economic, quick and valid method for counting foci in vitro micmmass cell cultures, and to provide a useful procedure relevant to this short-term test. MATERIAL AND METHODS： Midbrain cells and limb bud cells were collected from pregnant female rats at 13 days after conception and cultured constantly for 5 days in culture medium. The loci was counted by the new method introduced： the light microscopy field was divided into several compartments, moving across the compartments regularly and counting the numbers of foci. This paper also evaluated the accuracy and sensitivity of this procedure by testing reproducibility. RESULTS： The validity of test data was obtained via Grubbs test.The relative error of midbrain and limb bud cells were 0.776% and 0.213%; systerm standard deviations were 0.933 9 and 2.883 9; the coefficients of variation（CV） were 1.24% and 2.05%, respectively. CONCLUSION： High accuracy and sensitivity was obtained by the use of new parallel section mensuration method.The procedure could be used as aroutine method in micromass embryo cell ctdtures.