Dyes and stains

This information was collected from various online sources. It is for informational purposes only.

DAPI

• 358/461
• double stranded nucleic acid dye, associates with the minor groove of dsDNA, preferentially binds AT clusters
• stains both live and dead cells
• can be excited with a mercury-arc lamp or with the UV lines of the argon-ion laser
• better photostability than Hoesht dyes
• blue-fluorescent

Fluorescein diacetate

• fluorescein (494/518) is formed by intracellular hydrolysis of FDA
• bacteria can hydrolyze this
• leaks from cells
• fluorescein has a relatively high rate of photobleaching
• pH-sensitive fluorescence ref (pKa ~6.4) that is significantly reduced below pH 7
• relatively broad fluorescence emission spectrum (not good for multicolor use)

Carboxyfluorescein Diacetate

• fluorescein is formed by intracellular hydrolysis of CFDA
• carboxyfluorescein contains extra negative charges (structure) and is therefore better retained in cells
• been used in bacteria
• CFDA is moderately permeant to most cell membranes with uptake greater at pH 6.2 than at pH 7.4
• Oregon Green 488 (496/524) carboxylic acid diacetate is a more photostable version and less pH sensitive
• timing issues with how long you have to wait for the compound to be hydrolyzed ?

Propidium iodide

• 535/617
• nucleic acid dye
• preferentially stains dead cells
• excited by 488 laser
• red-fluorescent

SYTO

• from Molecular Probes now owned by Invitrogen
• nucleic acid dye
• membrane permeant
• series of blue-, green-, orange and red-fluorescent dyes
• says that better results are obtained in buffers without phosphate so no PBS?
• would require some optimization

Acridine Orange

• 500/526 for DNA, 460/650 for RNA
• nucleic acid binding dye
• cell-permeant
• emits green fluorescence when bound to dsDNA (525 nm) and red fluorescence when bound to ssDNA or RNA (650 nm).
• more common for eukaryotes?

Hoechst 33342

• 350/461
• double stranded DNA, minor groove–binding DNA stains
• cell-permeant nuclear counterstain
• excited with the UV spectral lines of the argon-ion laser and by most conventional fluorescence excitation sources
• emits blue fluorescence when bound to dsDNA.

SYBR Green I

• nucleic acid dye, better for DNA
• membrane permeant
• excited by 488 laser
• fluoresces green (maximum at 521 nm)
• tends to be used as a gel stain more often

SYBR Green II

• nucleic acid dye, better for RNA
• membrane permeant
• excited by 488 laser
• fluoresces green (maximum at 521 nm)
• tends to be used as a gel stain more often but also cited in flow cytometry