Chikamori K, Araki T, Sato K
Department of Chemistry, Naruto University of Education, Tokushima, Japan.
Acta Histochem. 1998 Feb;100(1):25-36. doi: 10.1016/s0065-1281(98)80004-4.
We measured in situ the activity of succinate dehydrogenase (SDH), one of the mitochondrial marker enzymes, in single Paramecium cells. SDH activity was detected with nitroblue tetrazolium (Nitro BT). Images of cells were captured every 30 sec at 590 nm, nearly the isosbestic wavelength of two reduction products of Nitro BT, by using a microphotometric system for image analysis. Activity was estimated by the slope of linear regression lines representing the relationship between total absorbance of the processed image and delta reaction time (real reaction time minus 30 sec). To investigate individual differences in Paramecium cell populations, SDH activity was measured in cells at various succinate concentrations. Paramecium SDH showed bimodal activity distribution patterns at three of four succinate concentrations tested. This result suggests that there are two groups of Paramecium populations with different SDH activity under control culture conditions. On the basis of the relationship between SDH activity and succinate concentration, mean Vmax and apparent Km values were estimated. A Km of 3.2 mM was found for Paramecium.
我们在单细胞草履虫中对线粒体标记酶之一的琥珀酸脱氢酶（SDH）的活性进行了原位测量。使用硝基蓝四唑（Nitro BT）检测SDH活性。利用用于图像分析的显微光度系统，在590 nm（接近Nitro BT两种还原产物的等吸收波长）处每隔30秒拍摄细胞图像。通过代表处理后图像的总吸光度与反应时间增量（实际反应时间减去30秒）之间关系的线性回归线斜率来估算活性。为了研究草履虫细胞群体中的个体差异，在不同琥珀酸浓度下测量细胞中的SDH活性。在所测试的四种琥珀酸浓度中的三种浓度下，草履虫SDH呈现双峰活性分布模式。这一结果表明，在对照培养条件下存在两组具有不同SDH活性的草履虫群体。根据SDH活性与琥珀酸浓度之间的关系，估算了平均Vmax和表观Km值。草履虫的Km值为3.2 mM。
