Hi!
Im trying to calculate ion strength and pH of solution.
Components of the solution:
47 ul of 1000 pmol DNA in Elution Buffer (10 mM Tris-HCl pH 8.5) was bound to 3 ul of 10 µM YoYo1 (flourescent dye) in DMSO and was mixed with 50 ul of Tris-EDTA buffer (100 mM Tris-HCl, 0.01 M EDTA pH 8.0).
About calculation of ion strength, Im confused what Ions are in the solution too.
Tris-HCl => Tris-H + Cl- ?
EDTA => Mg2+ + Ca2+??
DMSO => ??
Furthermore, how can I calculate it?
I think the pH range is about 8.0~8.5, because of two buffers.
Is there any method for calculating more exact pH value?
Thank you so much for your answers!
You could always measure the pH using pH paper and/or a probe. How precise do you need your measurement?
Dear Katie A Burnette
I cant get the pH probe. Is there any method for calculation? I think one decimal place is enough.
Assuming the buffers are correct, the pH is likely to be about 8.1 because the you have 100mM of Tris pH 8.0 and only 10mM of Tris at 8.5 (but I don't know if the YoYo has any buffering capacity and would change it).
Dear Michael J. Benedik
YoYo1(DMSO) doesnt have buffering capacity.
How can you calculated the 8.1 value?
Just an approximation. 100mM at 8.0 and 10mM at 8.5 should, if everything was linear, come out to 8.05, which rounds up to 8.1. I doubt the actual pH values are that precise to begin with, so it's a good approximation. You can never really be sure without measuring.
Do you have the solution made or is this just a calculation?
Honestly, I'd go the "empirical evidence" route and directly measure.
There are complicating factors that you don't have enough information to account for in your calculations.
Depending on your source of DNA and extraction method, the DNA will include proteins, ions, etc. Might even have lipids if it's a "dirty" prep. You can't assume that you have pure DNA.
DMSO is only very weakly acidic and will not deproteinate at pH 8-8.5. It's used as a solvent.
EDTA is a chelating agent and does not release ions. It's used to chelate Mg2+ and other metals to deactivate DNases.
Go ask for some pH paper (sometimes called litmus paper) or a pH probe.
Why don't you use a combined pH electrode to measure pH? It is the common type that you can find in any lab. You can calibrate it using two or more reference pH solutions. If you use five different pH references, you increase the accuracy of calibration. The common pHmeter uses only two solutions to calibrate. However, you can read the pH references in milivolts to build an external calibration curve with five pH reference solutions, if you will. Some pHmeters have the option of more than two references as well.
Michael J. Benedik
Thank you for your reply!
Ademario Junior Katie A Burnette
I just need a value by rough calculation, and I dont have enough amount of the solution (below 1ml) for the pH meter.
Thank you for your replies!
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