Imaging assays for microbial viability include reagents optimized for yeast and for bacteria as well as special formulations for biofilm research. As with mammalian cells, the assays rely on a combination of enzyme activity and membrane permeability to differentiate between live and dead cells. Life Technologies also offers microbial viability assays for analysis by flow cytometry.

Select assays for microbial viability

See also cell viability assays for flow cytometry
See also mammalian cell viability assays for imaging

Imaging Microbial Viability

  • Two-color assays for bacteria and yeast
  • Reduced biohazard option with fewer steps

Microbial viability can be determined using imaging assays that measure membrane permeability. A combination of membrane permeant and impermeant DNA dyes stains intact cells green and dead cells red. For bacteria, live cell determination can also be combined with fluorescent Gram staining.

For yeast cells, a membrane stain is used to detect both live and dead cells with a blue signal. In viable cells the vacuole stains orange/red, providing a two-color assay.

 
LIVE/DEAD® Yeast Viability KitStaining of yeast with the LIVE/DEAD® Yeast Viability Kit.

Imaging Biofilm Viability

  • Optimized for viability measurement in biofilms
  • Use esterase activity or membrane permeability

Measuring the viability of organisms growing in biofilm communities presents particular problems of cell penetration and accessibility. The FilmTracer™ Calcein dyes are non-fluorescent until esterases hydrolyze the acetoxymethyl ester, producing a highly fluorescent calcein. FilmTracer™ Calcein dyes are available in a range of wavelengths for multiplexing.

Two-color assays for bacterial viability using DNA binding dyes have also been optimized for use on biofilms. A combination of membrane permeant and impermeant DNA dyes stains intact cells green and dead cells red.

 
Bacterial Viability Kit

Staining streptomyces with the Bacterial Viability Kit.

Microbial viability can be determined using imaging assays to measure membrane permeability and DNA binding. For bacteria, live cell determination can also be combined with Gram staining.

 
Target Viability stains for bacteria Viability stain for yeast
Readout For live bacterial cells gram-positive cells stain green and gram-negative cells stain red Viability stains based on membrane permeability. Intact cells emit mostly green and dead cells emit mostly red Active cell vacuoles stain orange, live and dead cell walls stain blue
Fluorescent label SYTO® 9 SYTO® 9 SYTO® 10 FUN® 1
Hexidium iodide Promidium iodide Ethidium homodimer-2 Calcofluor White M2R
Standard filter set FITC FITC FITC FITC
Texas Red® Texas Red® Texas Red® DAPI
Ex/Em (nm) 480/500 480/500 484/505 488/560–610
480/625 536/617 535/624 365/440
Bibliography Citations Citations
Live cells Yes Yes Yes Yes
Fixable No  No No No
Platform Imaging Imaging Imaging Imaging
Format 1 kit/>1000 assays 1 kit/>1000 assays 1 kit/100 assays 1 kit
Cat. No. L7005 L7007 L7013 L7009

Microbial viability in biofilms can be determined using imaging assays to measure indicators of metabolic activity or esterase activity. Cell membranes increase their permeability in damaged and dying cells allowing the efflux of cellular components or the influx of dyes. Combination assays measure multiple parameters to provide more context for determining the status of both live and dead cells.

 
TargetBacterial viability stains compatible with and formulated for imaging biofilms
ReadoutViability stains based on membrane permeability. Intact cells emit mostly green and dead cells emit mostly red.The FilmTracer™ Calcein dyes are acetoxymethyl esters which serve as esterase substrates. They are cell permeant and become fluorescent after hydrolysis by esterases in viable cells. Viable cells exhibit fluorescence in a range of colors.
Fluorescent labelSYTO® 9

Calcein Orange

Calcein Violet

Calcein Green
Promidium iodide
Standard filter setFITC
Texas Red®

DAPI

FITC
Texas Red®
Ex/Em (nm)480/500
576/590

400/450

494/514
536/617
BibliographyCitationsCitations
Live cellsYesYesYesYes
FixableYesNoNoNo
PlatformImagingImagingImagingImaging
Format1 kit/>100 assays20 x 50 µg20 x 25 µg20 x 50 µg
Cat. No.L10316F10319F10320F10322