guanidinehydrochloride vs. guanidineisothiocyanate - Is there a difference concerning ability to denature proteins? (Nov/02/2006 )
Hi everybody,
I am supposed to do a denaturation of my protein in 6M guanidinhydrochloride followed by several wash steps in decreasing concentrations of guanidine hcl. the protein is in this case already fixated on nitrocellulose and is going to be subject to immunodetection afterwards.
now i found out that we do not have guanidine- hcl in the lab, but that we do have
guanidine iso thiocyanate.
So my question is : can i use that stuff , too to denature my protein? I guess each one of them is chaotropic, but is there a difference in using it with the nitrocellulose membrane? Or can i simply use urea for the same purpose?
Thank you so much,
Tobias
I am supposed to do a denaturation of my protein in 6M guanidinhydrochloride followed by several wash steps in decreasing concentrations of guanidine hcl. the protein is in this case already fixated on nitrocellulose and is going to be subject to immunodetection afterwards.
now i found out that we do not have guanidine- hcl in the lab, but that we do have
guanidine iso thiocyanate.
So my question is : can i use that stuff , too to denature my protein? I guess each one of them is chaotropic, but is there a difference in using it with the nitrocellulose membrane? Or can i simply use urea for the same purpose?
Thank you so much,
Tobias
more commonly used is potassium isothiocyanate; thiocyanate has a small hydration cover and, therefore, predominantly interacts directly with protein which is then denaturated; I suppose that you can take it for protein denaturation
In Gu Isothyocianate both ions are chaotropic, wheras in the HCl salt only guanidine is, so GuITC is a more powerful denaturant. It's also less soluble.
Use 4M GuITC instead of your 6M GuHCl
However I have no idea if you can use it with nitrocellulose membrane