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Dye exclusion artifact in flow cytometers.

Harald B Steen¹, Trond Stokke

¹Department of Biophysics, Institute for Cancer Research, Norwegian Radium Hospital, Montebello, Oslo N-0310, Norway.

|March 14, 2002

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Flow cytometry measurements can be affected by dye exclusion, where cells displace fluorescent dye solution. This artifact, caused by flow dynamics, impacts cell fluorescence signals, particularly at low flow rates.

Area of Science:

Biophysical techniques
Analytical chemistry
Cell biology

Background:

Cells displace fluorescent dye solution in flow cytometry, altering the baseline signal.
This dye exclusion phenomenon can significantly impact cellular fluorescence measurements under specific conditions.

Purpose of the Study:

To investigate the dye exclusion artifact in flow cytometry.
To understand how cell displacement of fluorescent solutions affects signal detection.

Main Methods:

Utilized the slit scan technique.
Studied monodisperse polystyrene particles in fluorescein solution to model the phenomenon.

Main Results:

Observed accumulation of dye solution before and after particles, with a void in between.

Related Experiment Videos

Identified flow constriction at the nozzle/flow chamber orifice as the likely cause, creating a fluorescence pulse.

Demonstrated that artifact magnitude is dependent on flow chamber design, dimensions, and sample flow rate.

Conclusions:

The dye exclusion artifact can influence flow cytometry measurements when cells are in highly fluorescent solutions.
This effect is more pronounced at low sample flow rates and depends on instrument geometry.