Journal
ANALYTICAL AND BIOANALYTICAL CHEMISTRY
Volume 395, Issue 5, Pages 1395-1401Publisher
SPRINGER HEIDELBERG
DOI: 10.1007/s00216-009-3119-8
Keywords
Epitope excision; Polystyrene beads; Prion protein; Antibody-antigen interaction; Mass spectrometry
Funding
- Swiss National Science Foundation [200020-124663]
Ask authors/readers for more resources
The binding epitope structure of a protein specifically recognized by an antibody provides key information to prevent and treat diseases with therapeutic antibodies and to develop antibody-based diagnostics. Epitope structures of antigens can be effectively identified by the proteolytic epitope excision-mass spectrometry (MS) method, which involves (1) immobilization of monoclonal or polyclonal antibodies, e.g., on N-hydroxysuccinimide-activated sepharose, (2) affinity binding of the antigen followed by limited proteolytic digestion of the immobilized immune complex, and (3) elution and mass spectrometric analysis of the remaining affinity-bound peptide(s). In the epitope analysis of recombinant cellular bovine prion protein (bPrP(C)) to a monoclonal antibody (mAb3E7), we found that epitope excision experiments resulted in extensive nonspecific binding of bPrP to a standard sepharose matrix employed. Here, we show that the use of amino-modified polystyrene beads with aldehyde functionality is an efficient alternative support for antibody immobilization, suitable for epitope excision-MS, with complete suppression of nonspecific bPrP binding.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available