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Characterization of phage peptide interaction with antibody using phage mediated immuno-PCR.
- Source :
-
Journal of immunological methods [J Immunol Methods] 2007 Sep 30; Vol. 326 (1-2), pp. 33-40. Date of Electronic Publication: 2007 Jul 18. - Publication Year :
- 2007
-
Abstract
- Real-time immuno-PCR (RT-IPCR) is a powerful technique that combines ELISA with the specificity and sensitivity of PCR. RT-IPCR of phage-displayed peptides exploits the unique physical associations between phenotype (the displayed peptide) and genotype (the encoding DNA) within the same phage particle. Previously, we identified phage peptides specific for recombinant antibodies (rAbs) prepared from clonally expanded plasma cells in multiple sclerosis (MS) cerebrospinal fluid (CSF) and subacute sclerosing panencephalitis (SSPE) brain. Herein, we applied phage-mediated RT-IPCR to study reactivity of these specific phage peptides for the rAbs. Compared to standard ELISA, which required greater than 10(4) or 10(5) phage particles to detect binding to rAbs, RT-IPCR detected binding with as few as 100 phage particles. RT-IPCR was also superior to ELISA in determining relative affinities of rAbs for phage peptides and was effective in screening MS CSF for IgG reactivity to phage peptides. Phage-mediated RT-IPCR is a rapid, high-throughput technology that avoids the requirement for synthetic peptides and will facilitate the identification of candidate peptides that react with the IgG in MS CSF.
- Subjects :
- Autoantibodies genetics
Bacteriophage M13 immunology
Enzyme-Linked Immunosorbent Assay
Humans
Immunoglobulin G analysis
Immunoglobulin G cerebrospinal fluid
Multiple Sclerosis cerebrospinal fluid
Multiple Sclerosis immunology
Peptides immunology
Recombinant Proteins genetics
Viral Proteins immunology
Autoantibodies metabolism
Bacteriophage M13 metabolism
Peptides metabolism
Polymerase Chain Reaction
Protein Interaction Mapping
Recombinant Proteins metabolism
Viral Proteins metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 0022-1759
- Volume :
- 326
- Issue :
- 1-2
- Database :
- MEDLINE
- Journal :
- Journal of immunological methods
- Publication Type :
- Academic Journal
- Accession number :
- 17669417
- Full Text :
- https://doi.org/10.1016/j.jim.2007.07.003