Studies on the reduction of 2,3,5-triphenyltetrazolium chloride as a viability assay for plant tissue cultures

Abstract

Optimum conditions for the reduction of 2,3,5-triphenyltetrazolium chloride (TTC) to a water-insoluble red formazan have been determined for suspension tissue cultures of Acer saccharum and Haplopappus gracilus. The standard incubation mixture is 0.8% TTC dissolved in a 2:1 solution of 0.05 M sodium phosphate buffer:Acer growth medium, pH 7.5. After the cells are incubated for 18–20 h in the dark at room temperature, the red formazan is extracted with 95% ethanol, and the absorbance is read at 485 nm. The use of the TTC procedure as a viability assay is supported by studies with Haplopappus tissue cultures, in which viability can be assayed by both plating in agar and the TTC procedure. These studies show that there is a quantitative correspondence between the decrease in the plating assay and in the TTC assay when these cells are subjected to freezing stress and plasmolysis stress.