Fluorescein Diacetate Staining

Detection of the viability of x-ray-exposed xylem cells was performed using a 9.6-μm fluorescein diacetate (Sigma-Aldrich) solution, in combination with fluorescence light microscopy. One plant was analyzed 10 d after first exposure to x-rays. Stem slices were obtained from the exposed part and 10 cm above this area. The stem was cut transversely, into 5-mm-thick slices, and immediately submerged into fluorescein diacetate solution for 30 min in the dark. Samples were rinsed with deionized water and placed onto a microscope glass slide. The sample surface was excited with green fluorescent light (λ = 490 nm) generated by a SOLA light engine (SE 5-LCR-VB; Lumencor) and observed for light at λ > 500 nm for the detection of living and metabolically active tissue (green signal) using an Axiozoom V16 macroscope (Zeiss) connected to an Axiocam 105 camera (Zeiss).