Good afternoon,
please how can you adjust pH of phage SM buffer? I presume that main ingredient that has influence of pH is Tris/HCl buffer, but according to literature its range of pH is from 7 to 9. But I have read articles when somebody has worked with SM buffer and its pH was 3 and 11. How is it possible? I`m confused. Please, have anybody experience with this problem? Thank you.
Hi dear Marie,
one time I was interested to isolate some phages against a bacterial strain, but I used the SM buffer that I found in all most articles with pH btw 7 and 7.5. maybe the articles that use SM buffer with pH 3 and 11 its for a specific kind of phages?
SM buffer (100 mM NaCl, 8 mM MgSO4, 50 mM Tris HCl (pH 7.5)) and stored at 4 °C .
SM buffer: 5.8 g NaCl, 2.0 g MgSO4·7H2O, 50 ml 1 M Tris-HCl pH 7.4, in 1 liter dH2O.
SM buffer [0.1 M NaCl, 8 mM MgSO4–7H2O, 50 mMTris–HCl, and 0.005% (wt/vol) glycerol) adjusted at neutral pH.
Marie Komárková - you can make the SM buffer to be whatever pH you want. Different phages might have different pH stability profiles. But in general most use a neutral pH in the 7-8 range. The actual buffer doesn't really matter, Tris is the most common, and some Mg ions (usually in the 5-10mM range of MgSO4 but can also be MgCl2). Obviously if you were working with phages of an acidophilic or alkaliphilic bacterium you might want to try a different pH range.
Dear Marie,
I think you may have stumbled onto a problem that sometimes researchers fail to appreciate. Buffers, but their very definition, are designed to alter the chemical environment of a solution by providing additional solutes which may be lacking or absorbed (e.g. Mg2+ which is absorbed by bacterial LPS or DNA) and 'forcing' or 'restricting' the pH of the solution to tolerable limits.
Buffers which are designed to control pH are only effective within a certain range of their pKa's - you can tweak the pH of your buffer with the addition of the appropriate acid or base when you make it, and if the buffering components are at sufficient concentration, they will be able to maintain that pH when you dilute and use the buffer with your sample (your sample will contribute to the over-all pH of the final solution and may contain strong buffering components or not).
You can keep adding acids and base to a buffer solution and force it along way from its pKa, but if you do this, it looses it's 'springiness' and it will not be able to maintain that pH when it is used - there is a danger that it will become more acidic or basic and the final equilibrium might be a long way away from what you wanted. Using a Tris buffer at pH 3 or 11 is an example of a very unstable system.
The solution to this is to use other chemicals for buffering including HEPES, MOPS, Acetate, etc, and different combinations of acids and bases allow a choice of optimal pKa's.
If you want to see how phage stability or infectivity varies with different buffers, then the best way forward is to re-make SM buffer, replacing Tris with your new acid-base combination. Alternatively, you could 'over-load' the Tris by adding a higher concentration of new buffer to SM, but this would of course dilute the other components of the SM buffer which may be important.
Regards, Andrew
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