Search Thermo Fisher Scientific
Search Thermo Fisher Scientific
The LIVE BacLight Bacterial Gram Stain Kit provides a novel one-step fluorescence assay for the determination of gram sign in living bacteria. When gram-negative and gram-positive bacteria are simultaneously stained with the membrane-permeant SYTO 9 dye and hexidium iodide; gram-negative bacteria fluoresce green and gram-positive bacteria fluoresce red.
1. Add 50 μL of late log-phase bacterial culture to 1 mL of filter-sterilized water |
2. Centrifuge for 5 minutes at 10,000 × g |
3. Wash the cells by removing the supernatant and resuspending the pellet in 1 mL of filter-sterilized water |
4. Prepare the cell staining solution by adding equal volumes of SYTO 9 (Component A) and hexidium iodide (Component B) to a microcentrifuge tube and mixing thoroughly |
5. Add 3 μL of the dye mixture per mL of bacterial suspension and mix thoroughly |
6. Incubate at room temperature in the dark for 15 minutes |
7. Put 5 µL of the stained bacterial suspension onto a glass slide and cover with a coverslip |
8. Image cells with the appropriate filters listed below |
SYTO 9 | Hexidium iodide | |
---|---|---|
Excitation/Emission | 480/500 nm | 518/600 nm |
Standard filter set | GFP | RFP |
Storage conditions | –20°C, protect from light | –20°C, protect from light |