9
4.
Count Apps
Brightfield App
The Brightfield app is used for quick, label-free total cell or particle counts. It is not used to determine cell
viability.
Trypan Blue App
Trypan blue is a cell membrane impermeable dye that only enters cells with compromised membranes.
Upon entry, the dye binds to intracellular proteins resulting in bluish colored cells. The trypan blue
exclusion assay enables a direct counting of live (unstained) and dead (blue) cells.
Published literature suggests that using trypan blue for samples with viabilities less than 70% may result
in higher reported cell viability values when compared to fluorescent-based detection methods.
Calculation:
% Viability =(Brightfield count / (Brightfield count+ Trypan Blue count)) *100.
Trypan Blue Concentration and Exposure Setting
Any brand of trypan dye can be used on the CellDrop but the concentration must be either 0.4%, which is
the common concentration of the dye sold by many vendors, or 0.2% trypan dye if the user chooses to
dilute the dye.
•
If the user is using 0.4% trypan dye then use the Trypan Normal Exposure setting
•
If the user is using 0.2% trypan dye then use the Trypan Low Exposure setting.
Trypan Blue Threshold
The brightfield channel captures a monochrome image so the darker cells stained with trypan have a
lower intensity. Therefore the trypan blue threshold setting is the intensity value from 1-100 at which
anything below is counted as a dead cell.
AO App
The AO App uses green fluorescence images to count live nucleated cells.
Acridine orange is a nuclear staining cationic fluorescent dye that enters live and dead cells and interacts
with DNA and RNA by intercalation or electrostatic attractions. When bound to DNA, it is spectrally very
similar to fluorescein, with an excitation maximum at 502 nm and an emission maximum at 525 nm.
Although AO enters dead cells, the intensity of the green fluorescence is very low for these cells
compared to the green fluoresce emitted from live cells. This app may be useful when counting solutions
containing either red blood cells and/or significant cellular debris.
The % cell viability is not provided when using this app.
PI App
The PI app uses brightfield images to capture live cell counts and red fluorescence images for counting
dead cells.
Propidium iodide (PI) is a red-fluorescent nuclear and chromosome stain that only enters dead cells.
Upon entry the PI stain binds to DNA by intercalating between the bases with little or no sequence
Summary of Contents for CellDrop
Page 1: ...rev MAR 27 2019 ...
