(by R.Jayakumar from roswellpark.org)
Wed Jan 4 09:24:33 EST 2012
Ideally the pH should not change much between 10X and 1X if deionised water was used for dilution. Presnce of NaCl has nothing to do with pH changes since it contributes neither hydroxyl nor hydrogen ions (unless otherwise it is contaminated with something else), the balance of which determines pH (not just ionic strength). The balance between the monobasic and dibasic is thus maintained the same whatever dilution the stock is diluted to. If stock concentrations affected pH, many companies that make 10X buffer stocks will be out of business. During dilutions make sure to use deionised water (Millipore MilliQ for example) or the ions in water can slightly affect the pH, but not that significantly as you observed.
From: methods-bounces from oat.bio.indiana.edu [mailto:methods-bounces from oat.bio.indiana.edu] On Behalf Of DK
Sent: Tuesday, January 03, 2012 6:08 PM
To: methods from magpie.bio.indiana.edu
Subject: Re: 10X PBS
In article <mailman.69.1325615292.3721.methods from net.bio.net>, sudheer sangeetham <sudheer.pbm07 from gmail.com> wrote:
>I have few doubts regarding phosphte buffer saline (PBS). Recently I found
>one article related to my work, in that they used PBS ( Nacl/iP ) pH-7.4.
>Nacl/iP does it mean, it should contain Na2HP04 and NaH2PO4 and Nacl but
>not KCl, am I right?
>I have prepared the buffer like this
>NaH2PO4 ( 0.038M)
>Na2HPO4.2H20 ( 0.162M)
>NaCl (1.5 M ) but when i check the pH it was showing 6.4, is it correct?
>did I do any mistake....
1. Measurng pH in ~ 2 M NaCl is not going to be very meaningful with
2. pKa is a function of ionic strenght - no surprise that 10X and 1X have
3. Dilute 10X, measure pH and if it's not above 7.2, you either made
an error making the solution or pH electrode is not measuring correctly.
4. The ratio you used above is for 0.2 M sodium phosphate, another
10X away from your 20 mM. (The difference between 200 and 20 will
be small but not negligible).
5. "PBS" can stand for many things. Cell culture PBS solutions
contain not ony some KCl (around 10 mM) but also some divalents,
CaCl2 and/or MgCl2.
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