Dyes and stains

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(Various dyes and stains that were researched for use in flow cytometry.)
(Dyes and stains)
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==Dyes and stains==
 
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This information was collected from various online sources.  It is for informational purposes only.
This information was collected from various online sources.  It is for informational purposes only.

Revision as of 13:21, 28 April 2005

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

Contents

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
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