Capasso Juan M, Cossío Belén R, Berl Tomás, Rivard Christopher J, Jiménez Carlos
Department of Renal Diseases and Hypertension, School of Medicine, University of Colorado Health Sciences Center, 4200 E. 9th Av., Denver, CO 80262, USA.
Biomol Eng. 2003 Jul;20(4-6):133-8. doi: 10.1016/s1389-0344(03)00037-6.
In this work, we propose the determination of cell viability in algal cultures by using a colorimetric assay widely used for estimation of cell proliferation in animal cell cultures. The method is based on in vivo reduction by metabolically active cells of a tetrazolium compound (MTS=3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenil)-2H-tetrazolium, inner salt) to a colored formazan, with maximal absorbance at 490 nm, that is released to the culture medium. For this purpose, we have tested two microalgae with high commercial value (Dunaliella and Spirulina) and two seaweeds with different morphology (Ulva and Gracilaria). Color development in this assay is directly proportional to the number of viable cells, to the incubation time in the presence of the assay solution, and to the incubation temperature. A direct significant correlation was found between algal photosynthesis rate and color development in all species used through this work. Moreover, the intensity of absorbance at 490 nm was significantly lower in stressed cells (e.g. in nutrient-limited cultures, in the presence of toxic substances, and in osmotically-stressed cultures). We conclude that cell viability of algal cultures can be rapidly and easily estimated through colorimetric determination of the reduction of MTS to formazan.
在本研究中,我们提出通过使用一种广泛用于评估动物细胞培养中细胞增殖的比色测定法来确定藻类培养物中的细胞活力。该方法基于代谢活跃的细胞在体内将四唑化合物(MTS = 3-(4,5-二甲基噻唑-2-基)-5-(3-羧基甲氧基苯基)-2-(4-磺基苯基)-2H-四唑,内盐)还原为有色甲臜,其在490nm处具有最大吸光度,并释放到培养基中。为此,我们测试了两种具有高商业价值的微藻(杜氏藻和螺旋藻)以及两种具有不同形态的海藻(石莼和江蓠)。该测定法中的显色与活细胞数量、在测定溶液存在下的孵育时间以及孵育温度直接成正比。在本研究中使用的所有物种中,藻类光合作用速率与显色之间发现了直接的显著相关性。此外,在应激细胞中(例如在营养受限的培养物中、在有毒物质存在下以及在渗透应激培养物中),490nm处的吸光度强度显著较低。我们得出结论,通过比色法测定MTS还原为甲臜,可以快速、轻松地估计藻类培养物的细胞活力。