Hello dears.
I have a question about buffers in protein works i hope that you will help me with this.
Why do we use a combination of buffers(it is better to say amphoteric compounds) and their salt in some buffer solutions for protein work? for example why do we use Buffers like PBS with this composition:
Na2HPO4 +KH2PO4+NaCl+KCl
or Sodium phosphate buffer with this composition:
Na2HPO4 +NaH2PO4+NaCl
Why we don't use just one buffer(amphoteric compound) and its salt for example buffer solutions with this compositon:
1.Na2HPO4 +NaCl
2.KH2PO4+KCl
3.NaH2PO4 +NaCl
4.K2HPO4+KCl
what is the differences between these two composition? is there any differences in their performance or their power?
is there any publications for details?
Thanks in advance.
A buffer will be just any or the four you say: Na2HPO4 , NaH2PO4, KH2PO4 , or K2HPO4. What buffers is the phosphate molecule though its speciation. Salts such as NaCl or KCl are just added to simulate the ionic strength found in your body, specially when dealing with cells as osmosis can occur and you do not want to stress the cells.
If you really understand the principle of a buffer system,you will know that buffer solution resist a change in pH(H+ or OH-) when acid or base is added.It consists of a conjugate acid and conjugate base not just any salt.However , we use buffer solution in protein purification with this property because these are essential components of all living organism.So, it required a physiological condition(pH) such as our blood to be buffered to maintain pH 7.4 to unchange metabolically-generated CO2. In protein purifcation,buffers are used to maintain stability and prevent loss of targeted protein with respect to pH and even temperature.The use of salt depends on the target protein and they are added when necessary to also improve stability and keep protein in solution by maintaining ionic strength of the medium.
I hope this will help.
All the best!
The mixture of monobasic and dibasic phosphate salts is used to adjust the pH of the phosphate buffer to the desired value between pH 6 and 8. For recipes, see this site:
http://cshprotocols.cshlp.org/content/2006/1/pdb.rec8543.full?text_only=true
Thank you dears for your suggestions.
According to your suggestions there is no difference between "(NaH2PO4 and KH2PO4)" and "( Na2HPO4 and K2HPO4)" buffering power i mean i can use either "(NaH2PO4 or KH2PO4)" as a monobasic and use either " (Na2HPO4 or K2HPO4)" as a dibasic. but each monobasic and dibasic supports the different buffering rang and in order to achieve a powerful and wide buffering rang we should use the mixture of these two buffers and their salts. i can even use one of the following compound according to my desired buffering rang:
1.Na2HPO4 +NaCl (for more basic rang)
2.KH2PO4+KCl (for more acidic rang)
3.NaH2PO4 +NaCl (for more acidic rang)
4.K2HPO4+KCl (for more basic rang)
.
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