Biostatus Limited - DRAQ5™ - LIVE cell comparison chart

DRAQ5™ LIVE cell comparison chart

The chart below compares six DNA dyes and their suitability for use in LIVE cell imaging applications:-

Nuclear Dyes → Technical Requirements↓	Prop. Iodide	TOTO3	TOPRO3	Hoechst 33258	DRAQ5	H2B.GFP
Practical
Membrane permeant ?	no	no	no	a)	yes	n/a
Fixation or permeabilisation required ?	yes	yes	yes	no	no	n/a
Stains intact primary culture cells ?	no	no	no	yes	yes	b)
Time (mins) / steps to achieve staining	60 / 5	60 / 5	60 / 5	5 / 1	5 / 1	n/a
Biological
Nucleolar / RNA staining ? c)	yes	yes	no	no	d)	n/a
Mitochondrial staining ? e)	no	no	no	no	no	no
Stoichiometric to Histone 2b ?	n/d	n/d	n/d	no	yes	n/a
Spatially quantitative to Histone 2b ?	n/d	n/d	n/d	no	yes	n/a
Phototoxic ?	n/d	n/d	n/d	f)	no	n/a
UV damage risk ?	n/d	n/d	n/d	g)	none	n/a
Pumped by MDR cells ?	n/d	n/d	n/d	h)	no	n/a
Spectral
UV ex. source required ?	no	no	no	g)	no	no
UV-induced "white-out" ? i)	no	no	no	g)	no	no
Argon / HeNe excitation ?	no	no	no	g)	yes	yes
Photobleaching (Hg lamp) ?	1 min	20-30s	5-10 s	~1 min	no bleaching	~1 min
Continuous imaging ?	n/d	n/d	n/d	n/d	>120 images	~ 60 images
2-photon excitation ?	n/d	n/d	n/d	f)	yes	n/a
Compatible with GFP, YFP, & RFP ?	some	yes	yes	no	yes	no

	Desirable quality for LIVE cell imaging
	Some desirable qualities for LIVE cell imaging
	Undesirable quality for LIVE cell imaging

a) DAPI is only semi-permeant

b) Can be difficult to transfect, hours before expression appears

c) SYTO dyes label both RNA & DNA in live & dead cells

d) Very weak nucleolar (RNA) staining

e) LDS-751 has been observed to label mitochondria

f) 2-photon excitation leads to rapid cell death

g) similarly DAPI

h) Multi Drug Resistant (MDR) cells may be ignored in cell screening

i) 10-15% of compounds screened in HTS/HCS assays will autofluoresce in UV, causing "white out" of the well under investigation.

For an independent review see:-

DNA Labeling in Living Cells. Martin et al., CytometryPart A 67A: 45-52 (2005).