测定细胞存活和增殖的MTT方法的建立

MTT方法具有简便,灵敏,稳定可靠,不需要同位素等优点。为了摸索测定细胞存活和增殖的MTT方法的各种最适条件,我们以Raji细胞为对象,比较了十二种不同的MTT甲(月替)溶解缓冲液。实验结果表明,20％SDS-50％DMF效果最好,溶解力强,溶解甲(月替)所需时间短,背景低,OD值高,不会引起培养基中小牛血清的絮状沉淀。其溶解的甲(月替)产物吸收峰在550—600nm之间。测定细胞范围为每孔500—400 000,这一范围能满足生物学和医学研究中多种测定的需要。

Use of MTT Assay for the Determination of Cell Viability and Proliferation

In order to optimize a rapid, sensitive and simple colorimetric cell viability assay (MTT assay), 12 different MTT formazan solubilization buffer have been tested for their ability to salubilize for-mazan as well as the activity to precipitate proteins in culture medium. It is found that the buffer consisting of 20% SDS-50% DMF(N,N-dimethyl formar.ide)is superior to the others. A absorbance peak of wavelenths 550-600nm is shown in the absorption spectra. All samples were measured with ELISA scanner at wavelenth 595nm for measurement and 655nm for reference. The linearity can be obtainedat cell numbers within the range of 500-400 000 cells per well and at serumm concentration less than 30%. MTT formazan is completely solubilized within 4 hours if the cells are not plated or centrifuged.