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Method Article
Molecular characterization of paired immunoglobulin heavy and light chains of clonally expanded antibodies in the presence of a strong polyclonal background
Birgit Obermeier
Institute of Clinical Neuroimmunology, Ludwig-Maximilians University, D-81377 Munich, Germany
Corresponding Author
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We describe a protocol for the direct molecular characterization of expanded antibody species detectable in minute amounts in human cerebrospinal fluid or other body fluids and tissues against a huge background of irrelevant antibodies. Antibodies are purified by affinity chromatography and applied to high resolution 2-dimensional PAGE under non-reducing conditions. Individual heterodimers consisting of heavy and light chains are isolated and analyzed by mass spectrometry. Since this rarely yields full sequence coverage, we reveal the complete sequences by alignment of the peptides to the immunoglobulin transcriptome of the same patient. Recombinant antibodies may then be produced and used for antigen searches or other experiments. We have analyzed antigens of oligoclonal band antibodies from the cerebrospinal fluid of patients with neurological inflammatory diseases, but the technology, which takes about two weeks, may be applied to many disease-relevant (auto)antibodies in human infectious and autoimmune diseases, as well as to B-cell malignancies.
Keywords
Oligoclonal band antibodies, B cells, Immunoglobulin, Proteomics, Transcriptomics, neurological inflammatory diseases
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Table 1 Overview on the properties of the different approaches to generate Ig transcriptome databases
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Associated Publications
Distinct oligoclonal band antibodies in multiple sclerosis recognize ubiquitous self-proteins
by Simone M. Brändle, Birgit Obermeier, Makbule Senel, +10
Proceedings Of The National Academy Of Sciences (06 September, 2017)
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This is a preprint. It has not completed peer review.
Method Article
Molecular characterization of paired immunoglobulin heavy and light chains of clonally expanded antibodies in the presence of a strong polyclonal background
Birgit Obermeier
Institute of Clinical Neuroimmunology, Ludwig-Maximilians University, D-81377 Munich, Germany
Corresponding Author
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 20 Oct, 2017
No community comments so far
Metrics
Comments: 0
HTML Views: ...
License:
This work is licensed under a CC BY 4.0 License. Read Full License
We describe a protocol for the direct molecular characterization of expanded antibody species detectable in minute amounts in human cerebrospinal fluid or other body fluids and tissues against a huge background of irrelevant antibodies. Antibodies are purified by affinity chromatography and applied to high resolution 2-dimensional PAGE under non-reducing conditions. Individual heterodimers consisting of heavy and light chains are isolated and analyzed by mass spectrometry. Since this rarely yields full sequence coverage, we reveal the complete sequences by alignment of the peptides to the immunoglobulin transcriptome of the same patient. Recombinant antibodies may then be produced and used for antigen searches or other experiments. We have analyzed antigens of oligoclonal band antibodies from the cerebrospinal fluid of patients with neurological inflammatory diseases, but the technology, which takes about two weeks, may be applied to many disease-relevant (auto)antibodies in human infectious and autoimmune diseases, as well as to B-cell malignancies.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
Associated Publications
Distinct oligoclonal band antibodies in multiple sclerosis recognize ubiquitous self-proteins
by Simone M. Brändle, Birgit Obermeier, Makbule Senel, +10
Proceedings Of The National Academy Of Sciences (06 September, 2017)