Electroelution of protein fromSDS-PAGE for use in immunization - (Nov/16/2000 )
Does anyone have any experience in removing a protein from an SDS-PAGE gel and using this to immunize a mouse to obtain polyclonal antisera? Any protocols would be helpful. Thanks.
You can cut the band of interest from the gel and mix it with CFA nicely. This can then be used for immunizatin. The boosters are given every 15 days in incomplete Freund's adjuvantGood Luck
Either do it Sunita's way or you can cut the band, put in a small bag of dialysis tubing with small amount of buffer and put the bag in a horizontal elctrophoresis apparatus and run low current-best done at ~20-30mA overnight at 4 C. Recover the protein solution and dialyse agianst PBS thoroughly and use. Or you can precipitate the protein with 5% ice chilled TCA.Cheers.
u can do in above two ways or stain the gel with 0.3M CuCl2, cut the unstained bands and destain with 0.25M tris and 0.25M EDTA
place the bands in a dialysis bag and dialyse against tris and glycine buffer
and then with pbs for immunization