Tris Hcl running gel buffer problems - (Nov/25/2008 )
I am trying to make 1.5 M Tris-Hcl pH8.8 and I dissolve the tris-hcl and the sds in water and it is clear and then I add NaOH to ph to 8.8 and it turned cloudy and after mixing for a while it is still cloudy. I followed the directions I don't know what to do. Any suggestions wouldbe appreciated. Thanks
What is SDS doing in 1.5M tris-HCl?
First make Tris, adjust your pH using HCl, then use it for any other solutions you need to make.
Check Tris-HCl Recipes here.
hth/
Edit: I just noticed that you are perhaps making SDS-PAGE gel upper chamber/SDS-glycine running buffer. For that you need to add HCl to bring the pH to 8.8, not NaOH. check these Recipes.
Another point of note is: Don't add hydroxide to Tris solutions, it destroys the buffering capacity of the Tris.
If you are making a Tris solution and it calls for Tris-HCl, just get some ordinary Tris and dissolve it, then adjust to the appropriate pH with HCl (this will take a lot of conc HCl for solutions near pH 7). For most applications actual Tris-HCl crystals are not needed.
Is it very cold in your lab? If you add SDS to your solution, this can fall out and the solutions gets blurry. Is the solution also cloudy without SDS?
In this case, I think the reason for cloudiness is obvious. NaOH.
No it is not cloudy when I do not add SDS. So if I am making 1.5M Tris-Hcl it should not have SDS right? That is what I thought but the recipe we have calls for 0.4 g of SDS. and 18.17 g of Tris.
you should add the sds after you adjust the pH of the tris.