please, i want to make precipitation of protein samples using ammonium sulfate 40% how can i calculate it as i read about this method and see more tables regarding temp. which can not able to understand ? and which is more preferable using sat. ammonium sulfate sol. or addition of solid particles to the sample ?
Http://encorbio.com/protocols/AM-SO4.htm
please see the above link to help you calculate for protein precipitation with ammonium sulphate.
i hope this helps.
Dear Dina,
You would use an ammonium sulfate solution of desired percentage to be used to precipitate a protein at 0°C, as described by Green & Hughes (1955). The following calculator would be helpful.
http://www.proteinchemist.com/cgi-bin/s2.pl
http://kuchem.kyoto-u.ac.jp/seika/shiraishi/protocols/as_precipitation.html
Precipitation of proteins is performed at about 0°C to avoid possible denaturation of proteins even though the % saturation values shown in the table are based on the values at 25°C described by Green and Hughes {Methods Enzymol. 1, 67-90 (1955)}.
http://proteincrystallography.org/protein-purification/ammonium-sulphate-table.php Pick up your initial concentration of ammonium sulfate in percent from saturation from the left column, then select the required final concentration of ammonium sulfate in percent from saturation from the top row. Intersection gives you and exact amount of ammonium sulfate in grams for 1 L of initial solution.
http://www.biovision.com/manuals/7096.pdf
Ammonium sulfate is employed as a step in the fractionation (and initial purification) of a protein by "salting out". At higher salt concentrations, most proteins display less solubility, and this is called salting out; this is utilized to fractionate proteins. Concentration of the salt required to precipitate proteins varies from one protein to another. Salting out can be useful in the concentration of a dilute solution of a protein; salt can be removed by dialysis or gel filtration.
Green, A. A., and Hughes, W. L., Protein fractionation on the basis of solubility in aqueous solutions of salts and organic solvents. Methods Enzymol., 1, 67-90 (1955).
Deutscher, Murray P., Methods in Enzymology, vol. 182, Guide to Protein Purification, New York: Academic Press, 1990. 285-306.
Best...
Hi
I used this site formula for calculating and it works.
http://tmw.wzw.tum.de/fileadmin/user_upload/Methodendatenbank/Dateien/AS-Calculator.html
On predicting the pH of a pure (NH4)2SO4 aq. sol.; see: https://www.researchgate.net/post/What-is-the-difference-between-using-ammonium-sulfate-and-using-sodium-sulphate-in-protein-ppt-and-seperation
I hope these documents will be helpful to you to do the protein precipitation at 0 C and 20 C
I use this link to calculate the amount of ammonium sulfate to precipitate protein.
https://www.encorbio.com/protocols/AM-SO4.htm
- Badges
- Science method