Fig. 2.

Example of Live/Dead staining kits applied to two bacterial samples and a eukaryotic sample. (A) A pure culture of E. coli was grown in LB medium overnight at 37 °C to an OD₆₆₀ of 0.4. The cells were incubated with 100 mM H₂O₂ for 1 h at 37 °C. The sample was then stained with the LIVE/DEAD BacLight Bacterial Kit-L-7007 (Invitrogen, Grand Island, NY, USA) for microscopy according to the manufacturer’s instructions. A 10-μL aliquot was examined by fluorescence microscopy on a Carl Zeiss Axioskop using a filter with an excitation 488 nm and emission 528 nm. The live cells fluoresce green. (B) A developing biofilm on a glass slide created by incubating the slide in a solution containing three bacterial species: (1) Serratia marcescens ATCC 14756, (2) Corynebacterium xerosis ATCC 373, and (3) Staphylococcus epidermis ATCC 14990. It was stained using the LIVE/DEAD BacLight bacterial viability kit (PI/SYTO) [Molecular Probes]. Here, the live cells fluoresce green while the dead cells fluoresce red. (C) Yeast cells were stained with the LIVE/DEAD Yeast Viability Kit L-7009 (FUN 1 cell stain). The yeast were grown overnight in Sabouraud medium at 28 °C and then incubated with 100 μM H₂O₂ for 1 h. The samples were stained with FUN 1 cell stain according to the manufacturer’s instructions. The cells (10 μl aliquots) were viewed under a fluorescence microscope Axioskop (Carl Zeiss) with an excitation 489 nm and emission 539 nm. In contrast to the images of bacteria, here, the live cells form red fluorescent structures, while the dead cells are distinguished by a diffuse, green fluorescence. E. coli and S. cerevisiae micrographs were obtained by coauthor Balk, and the mixed bacterial micrograph was obtained by coauthors Adams and Lymperopoulou