GST fused protein as immunogen?

Zhonglin Chai Zhonglin.Chai at baker.edu.au
Thu Jun 14 03:59:54 EST 2001


Dear Jinguo,

It seems that the antiserum you raised in rabbit contains very low titer, or none, of Abs to your protein, although high titer of Abs to GST is raised. GST is always a very strong immunogen that stimulates a strong immune response with high titer of Abs. But you don't know whether the fusion partnre of the protein of interest is immungenic, some of which are very poor immunogen that can hardly raise enough Abs. You may test the serum for the Abs to your protein by immunoblotting an endogenous protein (cell lysates, for example) or fusion protein with other partner than GST (maltose binding protein, for example) to confirm your serum is good in the first place. If the serum contains Abs to both GST and your protein, absorption with GST column to remove Abs to GST, followed by affinity purification of the Abs to your protein on a fusion protein column (GST fusion protein would do) should be good enough to give you highly purified Abs for applicaiton.  

Hope this of help.

Zhonglin Chai

Zhonglin Chai, PhD

Hypertension Laboratory
Baker Medical Research Institute
P.O. Box 6492
Melbourne 8008
Victoria, Australia
Telephone: +61 3, 9522 4357
mobile : 0413 58 1940, or international +61, 413 58 1940
Fax: +61 3, 9521 1362 
email: zhonglin.chai at baker.edu.au


>>> Jinguo Wang <n84jw at morgan.ucs.mun.ca> 06/13/01 04:48AM >>>
Anyone has experiences with GST fused protein as immunogen? My
recombinant GST fused protein is in inclusion body form and the rabbit 
got response against the fused whole protein. However, I have some
trouble with separating the abs against the GST and my real target
protein. After absorption with the rGST, it seems I lost everything..

Can someone please give me any suggestions? 

Thanks in advance.

Jinguo




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