Top-down proteomics analysis of the antigen binding region of IgG to detect antigen-specific sequences that are shared between individuals
EMSL Project ID
43090
Abstract
Autoantibodies play an important role in the diagnosis of cancer and autoimmune diseases and are increasingly used as biomarkers in the early detection of cancer. Disease specific antibodies, including autoantibodies, are generally detected by their binding to specific antigens. We propose to use an alternative approach, namely to directly detect antibodies against autoantigens and tumor specific antigens by the sequence of their complementarity determining regions (CDRs) using mass spectrometry (MS) sequencing, and to use these CDRs as biomarkers. This approach is valid when individuals produce antibodies against a particular antigen that have CDR sequences in common. Experimental data (including our own data) on tryptic digests of IgG from patient samples and from an animal model support this hypothesis of convergence in antibody sequences. However, a disadvantage of experiments using a digestion step is the loss of the positional information of peptides in the IgG molecule. The high diversity in the IgG molecules, especially in the CDRs, makes it difficult or error-prone to determine the location on the IgG backbone based on homology. In addition it is impossible to reconstruct the sequence of complete IgGs from data obtained from a bottom-up approach. Therefore, top-down measurements with high resolution FT equipment, which can deliver data of immunoglobulin segments that span the entire variable domain, is sought. Herein, we propose to use a combination of a bottom-up and top-down approach on EMSL’s high magnetic field FT-ICR MS to perform a proof-of-principle experiment.
Project Details
Project type
Limited Scope
Start Date
2011-01-07
End Date
2011-03-09
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members