The LIVE/DEAD BacLight Bacterial Viability Kits utilize mixtures of our SYTO™ 9 green-fluorescent nucleic acid stain and the red-fluorescent nucleic acid stain, propidium iodide. These stains differ both in their spectral characteristics and in their ability to penetrate healthy bacterial cells. When used alone, the SYTO 9 stain generally labels all bacteria in a population — those with intact membranes and those with damaged membranes. In contrast, propidium iodide penetrates only bacteria with damaged membranes, causing a reduction in the SYTO 9 stain fluorescence when both dyes are present.

The LIVE/DEAD Bac Light Bacterial Viability Kits have been thoroughly tested with a variety of organisms and under several conditions. The kits are ideal for fluorescence microscopy as the live (green fluorescent) and dead (red fluorescent) cells may be viewed separately. They are also suitable for quantitative analysis with a fluorometer, fluorescence microplate reader, flow cytometer, or other instrumentation.

Traditional methods of assessing bacterial viability rely on metabolic characteristics or membrane integrity. However, due to differences in morphology, cytology, and physiology among the many bacterial genera, developing a universally applicable direct-count viability assay has been challenging. The LIVE/DEAD BacLight Bacterial Viability Kits based on membrane integrity allow researchers to distinguish live and dead bacteria easily, reliably, and quantitatively in minutes, even in a mixed population with diverse bacterial types.

The LIVE/DEAD BacLight Bacterial Viability Kit is available in three different formats:

LIVE/DEAD BacLight Bacterial Viability Kit, for microscopy (Cat. No. L7007)
The LIVE/DEAD BacLight Bacterial Viability Kit, for microscopy includes two different premixed solutions of the SYTO 9 and propidium iodide (PI) in 300 μL of DMSO. The fluorescent dyes are mixed at different proportions in each one: Component A – 1.67 mM SYTO 9 & 1.67 mM PI, Component B – 1.67 mM STYO 9 & 18.3 mM PI. This kit is available for customers who have already developed protocols using the premixed dye formulation.

LIVE/DEAD BacLight Bacterial Viability Kit, for microscopy & quantitative assays (Cat. No. L7012)
The LIVE/DEAD BacLight Bacterial Viability Kit, for microscopy & quantitative assays includes two separate solutions of the SYTO 9 (3.34 mM) and propidium iodide (20 mM) dyes in 300 μL of DMSO. The separate staining components enables flexibility for optimization and facilitates the calibration of bacterial fluorescence for quantitative procedures.

LIVE/DEAD BacLight Bacterial Viability Kit (Cat. No. L13152)
For added convenience, the LIVE/DEAD BacLight Bacterial Viability Kit contains 10 pairs of the premeasured separate SYTO 9 and propidium iodide dyes in sealed polyethylene transfer pipets. In addition to the convenient packaging in the sealed pipets, the fluorescent dyes are supplied dry, without DMSO or other solvents. This formulation does not require refrigeration and remains stable at temperatures ≤–20°C once reconstituted.

All three kits come with 10mL of BacLight mounting oil for bacteria immobilized on membranes. The refractive index at 25°C is 1.517 ± 0.003.

Any physiological buffer between pH 7.0–8.0, including PBS, can be used to dilute the SYTO dyes for the staining solution.

At the recommended reagent dilutions and volumes, kits L7007 and L7012 contain sufficient material to perform ≥1000 individual tests in 96-well assay plates, many more tests by fluorescence microscopy or ~200 tests by flow cytometry.

In kit L13152, each applicator pair contains sufficient dye to perform 50 individual tests in a 96-well assay plate, ~1000 assays by fluorescence microscopy or 10 tests by flow cytometry.

The LIVE/DEAD BacLight Bacterial Viability Kits have been tested on the following bacterial species: Bacillus cereus, B. subtilis, Clostridium perfringens, Escherichia coli, Klebsiella pneumoniae, Micrococcus luteus, Mycobacterium phlei, Pseudomonas aeruginosa, P. syringae, Salmonella oranienburg, Serratia marcescens, Shigella sonnei, Staphylococcus aureus and Streptococcus pyogenes. All these bacterial types have shown a good correlation between the results obtained with the LIVE/DEAD BacLight Bacterial Viability Kits and those obtained with standard plate counts.

These tests were performed on logarithmically growing cultures of organisms. In addition, we have received favorable reports from researchers who have used these kits with: Agrobacterium tumefaciens, Edwardsiella ictaluri, Eurioplasma eurilytica, Lactobacillus sp., Mycoplasma hominus, Propionibacterium sp., Proteus mirabilis and Zymomonas sp.