Hi everyone,
I got into the crystal violet staining assay for determination of cell viability recently.
As far as I know, CV stains the DNA in the adhered live cells and O.D. can be measured to determine the viability?
Why can this work?
Is it because the more viable cells contain more DNA which is to be stained while dead/semi-dead cells contain less DNA?
How does this method distinguish different cells within the adherent surface?
dear friend
I read the protocole and I think you don't consider Washing step which had been done prior to violet crystals formation. So, when you go through washing step, clearly you eliminate all dead (when an adherent cell dying, it detached from plate) cells in which formed violet colour just will be the representative of proliferative and alive cells.
And Don't forget that, OD magnitude is a function of number of alive cells (each cell has clear amount of DNA and Proteins).
I hope this could help you.
Amir
Kaushiki Chatterjee Amir Jafari Subhash C. Juneja
Hi everyone!
Thanks for the detailed explanation!
I understand I would be washing away all the dead cells which are not adherent.
But for the rest of adherent cells on the substrate, how could the cell viability to be determined? Like portion of the cells are more active which can be measured due to more DNA while the rest of the cells are less active where in this case the DNA amount would be smaller?
Or does the Crystal Violet stain binds indifferently to all the cells and the result well are kind of uniform?
Kaushiki Chatterjee
Hi Kaushiki,
So that means the remaining adherent cells will adsorb the stain indifferently and the OD is only used to distinguish how many live cells are there.
I mean, that relies on the assumption that all the dead cells are removed and it was trying to count how many cells are left on the surface, as sort of indicator of viability.
Is that right?
MTT assay is a good indicator of viability.We used it to determine the ameliorative effect of Oroxylum extract on lymphocytes subjected to toxic effects of bidi tobacco extracts
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