Can nucleic acid stain distinguish "virion" and "free viral nucleic acid"?

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Engelbert Buxbaum

I suppose this might work for enveloped virus, if you use a stain that is not membrane-permeable.

Zaizhi Yu

Mr. Buxbaum, thank you for your kind advice. That makes sense. I am targeting non-enveloped viruses now, so I will go find other solutions

Michael J. Benedik

You might be able to remove any free DNA from the sample either by nuclease treatment or some absorption step and then use a stain to detect virions. You won't know for certain whether it will work or not until you do some appropriate controlled and reconstruction experiments.

If this is a test you intend to perform regularly then it might be worth the time to develop antibodies against your virions (assuming it is a unique virus and not a population of different viruses).

Thank you for the very detailed advice.

I have heard that free DNA and DNA in virions can be distinguished just by the staining, but I have a doubt on that.

The pretreatment completely makes sense, and I will follow that.

I have got one more question. Is it popular to stain viral RNA?

Perikles Simon

I am following this question because of the waste water aspect. Both, DNA and RNA, can also be present in vesicles released from human and animal cells into waste water. The content of such vesicles (exosomes and microparticles) will also be protected from nuclease treatements (just like the DNA and RNA of all microbes). So in the end, if there is no specific dye penetrating viral capsides only, which I am not aware of; you will need a kind of specific detection approach to make sure that you are indeed measuring viral RNA or DNA. But first, you may want to check Molecular Probes products, since they offer all different kinds of specific dyes (mitotracker and many other).

Sibnarayan Datta

Dear Dr. Yu,

you can perform Epifluorescent microscopy using SYBR Gold dye to see and enumerate virus particles under a fluorescent microscope. For details, you may refer to this excellent article:

https://www.nature.com/articles/nprot.2009.10

best wishes,