Measuring freezing tolerance: electrolyte leakage and chlorophyll fluorescence assays.

Quantitative assessment of freezing tolerance is essential to unravel plant adaptations to cold temperatures. Not only the survival of whole plants but also impairment of detached leaves after a freeze-thaw cycle can be used to accurately quantify plant freezing tolerance in terms of LT50 values. Here we describe two methods to determine the freezing tolerance of detached leaves using different physiological parameters. Firstly, we illustrate how to assess the integrity of (predominantly) the plasma membrane during freezing using an electrolyte leakage assay. Secondly, we provide a chlorophyll fluorescence imaging protocol to determine the freezing tolerance of the photosynthetic apparatus.