Protein isolation and lack of sop's - (Feb/24/2013 )

Hello


We are a group of university students from leiden (The netherlands )
We are all Microbiologists so whe do know very few about proteomics.
we have 1 bacteriologist 1 mycologist and all round microbiologist.
and i'm the medical microbiologist and biochemist


we have to do an assigement to purify the enzyme peroxidase out of horse radish.
for this experiment we need several SOP's

1) extraction ( SOP made by myself)
2)Determination of the RZ Value for PEROXIDASE (we have a sop but it's not good ) can anyone help us find a better one?
3) BCA analysis i used that sop form santa cruiz university
4) peroxidase test we have one dated 1994 are there any recent sop's?
5)Ammonium sulfate precipitation whe have an UAS Leiden Sop for that but ofcourse it's shady and makes no sense.

6)dialysis we have that found already
7)Concanavalin-A chromatography - can any one help us with that ? we can't find
8) Gel filtration with TSK2000 - we have to use this technique we have got very old lecture stuff about it but it's not that good.
i know about Size exclusion but not what TSK2000 is so please anyone can you help 3 students out?

9) the last SPE-gel electrophoreses and subsequent DAB staining of the peroxidase isozymes
we have that done before i found half of that protocol because UAS Leiden gave us that before.




any hints ,tips or links to good SOP's we will be very happy

so please help us out

with kind regards
Timo
student :Medical microbiology
University of Appleid Science Leiden

you can download handbooks on affinity chromatography (including con-a) and gel filtration from this webpage:

ge life sciences

you can find an assay for hrp and links to more information at this page:

worthington

the attached ms word document will give more information on precipitation with ammonium sulfate.

tsk2000 is a specific gel filtration column for use on hplc apparatus. tsk is the company and 2000 is the packing type. this page will download the tsk brochure on sec:

brochure

hope these help.