I performed homogenization of pancreas tissue for protein extraction by using 1X PBS(phosphate buffer saline)at physiological PH,I transferred the tissue from a tube by old forceps,I notice that particle of Fe stuck on tissue afterward with homogenization and centrifugation I noticed that the protein solution became red color.My question:Is this color normal because of tissue?and in case if its Fe is there any effect of protein if I want to separate them on SDS-PAGE and then cut and digest to do mass spec?
Iron ions can catalyze oxidation, although most of the iron oxide rust will be insoluble. Put some EDTA in the buffer to chelate any soluble iron. If at some point you centrifuge the sample, insoluble rust will be in the pellet.
I don't think it will effect the separation on sds page. The iron will get separated from your protein while running on sds page unless bound tightly. As you mentioned you have already centrifuged the samples which has removed insoluble fe, now you can do dialysis with mwco of membrane less than that of your interested protein to remove soluble fe.
You might want to check databases online if your particular target protein has some iron-binding properties. If not, you don't have to worry about that.
- Badges
- Science topic
- Similar topics
- Chemistry
- Biochemistry