A colleague of mine used ammonium sulfate precepitation for enzyme purification. Chapter 4.6 in following link (hope this kind of long link will works).

https://www.google.de/url?sa=t&rct=j&q=&esrc=s&source=web&cd=1&cad=rja&uact=8&ved=0ahUKEwjwtJK_zavJAhXDqA4KHfeZAKYQFggmMAA&url=http%3A%2F%2Frzbl04.biblio.etc.tu-bs.de%3A8080%2Fdocportal%2Fservlets%2FMCRFileNodeServlet%2FDocPortal_derivate_00036658%2FDiss_Zein_Marwan.pdf%3Bjsessionid%3D83BD4DB853DB204F8CAD1F7BCEF644B0&usg=AFQjCNGg5-kp7PgeB4KKjSCukitZpgj9hg

Otherwise google his thesis ("Integration of Reaction and Product purification for

trienzymatic catalyzed synthesis of Laminaribiose from Sucrose")

Thanks Thomas!

Also take into account that the resolution of ammonium sulfate precipitation is really low, hence most people use it at the beginning, rather than the end of their purification schemes (unless they want to simply concentrate their preparations). It is better to use it before, not after an affinity purification step.

Hi Alejandro, have you done purification using ammonium sulfate before? Mind to share how you did it? if I plan to optimize the concentration to purify out those remaining impurities only after affinity purification, do you think I can do that? There is a lot of impurities in the crude and i think it's quite hard to precipitate it out. Don't you think that?

Hi Alex, I used quite a lot of ammonium sulfate in my youth, PhD and shortly afterwards. If you do not have any clue on at which saturation your protein precipitates, you may like to run some preliminary assays. The idea is to know within which saturation percentages your protein precipitates. Before reaching the inferior % range other proteins will precipitate but not your protein. After reaching the superior % range your protein will precipitate and other proteins may still be in the "supernatant".

In any case remember that ammonium sulfate is just a step in  a purification procedure, it serves to eliminate undesired proteins plus concentrate the sample by centrifugation and resuspension in small volumes.

Remember also that , at the end, if you have your protein precipitated with ammonium sulfate you may need to get rid of the ammonium sulfate that may affect further purification steps or functional assays.

Best

Yes, I've done it before. First you add ammonium sulfate at a specific saturation % to precipitate proteins other than your target protein, then add additional ammonium sulfate at a higher saturation % to precipitate your protein while -hopefully- leaving other contaminants behind (provided that the protein does not crash irreversibly in this step). The exact % have to be determined experimentally beforehand; go google a bit and you'll find better descriptions than anything I can provide.

The point is -and judging from your comments, you are well aware of this- that AS precipitation is not very selective. You use it in a crude lysate and it gets rid of a lot of contaminants, but will never yield a 'pure' protein by any sensible criterion, and the reason is that the minimum AS saturation % at which all of a given species will precipitate is not a single well-defined value, but a range that often overlaps with that of other protein species. If you use it after an affinity step it may improve the purity of the resulting preparation, but will never work as well as another run on -say- a gel filtration column.