How does the RBC lysis buffer work?
Posted 04 November 2010 - 05:43 AM
I used this recipe:
1 l ddH20
8.26 g Ammonium Chloride (NH4Cl)
1 g Potassium Bicarbonate (KHCO3)
0.037 g EDTA
The rumor has it that it has something to do with the Na+/K+ATPase activity, but this isn't as detailed as I would like to know how it works.
I would very much appreciate an explanation how this buffer work, or maybe a hint to where I can find litterature, or the like, on this.
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Posted 11 November 2010 - 11:00 PM
Posted 11 November 2010 - 11:43 PM
Posted 12 November 2010 - 12:08 AM
The other option is that many of these 80 post viewers spent some time looking into this for you (like myself, who spent over an hour on the internet trying to find out) and couldn't find anything more detailed than what you already know..........
Well that is also possible I guess. I apologize to all of you nice people who tried to help me out, and I also thank you for your time and effort!
Posted 23 July 2013 - 12:27 PM
Posted 25 July 2013 - 02:17 AM
Please refer to the reference below:-
The important bit is:-
Mammalian erythrocytes placed into distilled water
will swell until at some critical point the rising
pressure in the cell interior overwhelms the tensile
strength of the cell membrane and hemoglobin escapes
(hemolysis). Delano (1995) developed a
model to predict the critical set of physical conditions
that result in osmotic rupture of mammalian
erythrocytes. The model determined that the
fate of the erythrocyte in a hypotonic solution depends
on tonicity, the initial sphericity of the cell,
the initial fraction of its volume not occupied by
water, and the maximum fractional amount by
which the surface area can be stretched.
We have been using your lysis buffer (NH4CL) to remove red blood cells from Ficoll-Paque isolation of whole blood. The RBC's are an unwanted contaminant as we are after Neutrophils, Monocytes, Platelets etc. By using this lysis buffer these cells are not activated in any way as to ruin our experiments i.e. degranulation and chemotaxis of Neutraphils.
I hope this gives you some more understanding of what is involved in the hypotonic lysis of RBC's
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