Specificity of recombinant antibodies generated from multiple sclerosis cerebrospinal fluid probed with a random peptide library

Yu, Xiaoli; Gilden, Donald H.; Ritchie, Alanna M.; Burgoon, Mark P.; Keays, Kathryne M.; Owens, Gregory P.

doi:10.1016/j.jneuroim.2005.11.010

Cited by 39 publications

(53 citation statements)

References 39 publications

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“…Furthermore, one study analyzing recombinant scFV fragments constructed from CSF B cells found an axon-directed antibody response [32]. Another recent study, applying random peptide libraries, failed to identify potential target protein antigens of MS-mAb [33] while others were able to demonstrate MBP reactivity of clonally expanded CSF B cells by means of recombinant Fab fragments [34]. In the current study, we reconstructed antibodies produced by cePC in order to determine whether they display patterns of tissue reactivity that would enable us to understand their potential involvement in the immunopathogenesis of MS.…”

Section: Discussionmentioning

confidence: 99%

Clonally expanded plasma cells in the cerebrospinal fluid of MS patients produce myelin‐specific antibodies

et al. 2008

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Clonally expanded plasma cells (cePC) and their presumed products, oligoclonal immunoglobulin G bands (OCB), are characteristic findings in the cerebrospinal fluid (CSF) of patients with multiple sclerosis (MS). While cePC and OCB strongly suggest an involvement of B cell-dependent immune mechanisms in the pathogenesis of MS, their actual pathological relevance and target antigens remain unknown. To further understand the potential role played by cePC, we generated a panel of monoclonal antibodies (MS-mAb) from CSF-derived cePC from four patients with early or definite MS. Single-cell RT-PCR of correctly paired heavy and light chain immunoglobulin genes from individual cePC ensured the subsequent resurrection of their original antigen specificity. Immunofluorescence stainings of MS lesion tissue with MS-mAb revealed myelin reactivity in the cePC repertoire of all four patients and intracellular filament reactivity in one patient. While myelin staining by MS-mAb was only rarely detectable in non-MS CNS white matter tissue, it was greatly enhanced at the edge of demyelinating lesions in MS brain tissue. Our findings provide conclusive evidence for the presence of an antigen-driven B cell response in the CSF of MS patients directed against epitopes present in areas of myelin degradation.

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Section: Discussionmentioning

confidence: 99%

Clonally expanded plasma cells in the cerebrospinal fluid of MS patients produce myelin‐specific antibodies

et al. 2008

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“…In a recent publication about the specificity of recombinant antibodies cloned from the CSF fluid of two MS patients, several peptide sequences were identified, which specifically bound to the antibody clones. However, no homologies to known human or viral proteins were identified (Yu et al, 2006). The data summarized here mark our first analysis of the protein antigen specificity of clonally expanded CSF B cells isolated from patients with RRMS.…”

Section: Discussionmentioning

confidence: 85%

Antigen specificity of clonally expanded and receptor edited cerebrospinal fluid B cells from patients with relapsing remitting MS

Lambracht-Washington

O’Connor

Cameron

et al. 2007

Journal of Neuroimmunology

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We re-engineered the immunoglobulin rearrangements from clonally expanded CSF B cells of three Multiple Sclerosis patients as Fab fragments, and used three methods to test for their Ag-specificity. Nine out of ten Fab fragments were reactive to Myelin Basic Protein (MBP). The one Fab that did not react to MBP was a product of receptor editing. Two of the nine MBP-reactive Fabs were also reactive to GFAP and CNPase, indicating that these clones were polyreactive. Targeting the mechanisms that allows these self-reactive B cells to reside in the CSF of MS patients may prove to be a potent immunotherapeutic strategy.

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“…Full-length human IgG1 rAbs were produced in mammalian tissue culture cells (Invitrogen, Carlsbad, Calif.) from V region sequences of expanded SSPE brain plasma cell clones as described previously (3). Briefly, V H regions were cloned into pIgG1Flag, a modified version of the episomal expression vector pCEP4 (Invitrogen) that contains a full-length human IgG1 C region domain and a C-terminal Flag epitope (21). Amplified full-length light-chain (L-chain) sequences were cloned independently into the unmodified pCEP4 vector.…”

Section: Methodsmentioning

confidence: 99%

Screening Random Peptide Libraries with Subacute Sclerosing PanencephalitisBrain-Derived Recombinant Antibodies Identifies Multiple Epitopes in the C-Terminal Region of the Measles Virus Nucleocapsid Protein

Owens¹,

Shearer²,

Yu³

et al. 2006

J Virol

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Infectious and inflammatory diseases of the CNS are often characterized by a robust B-cell response that manifests as increased intrathecal immunoglobulin G (IgG) synthesis and the presence of oligoclonal bands. We previously used laser capture microdissection and single-cell PCR to analyze the IgG variable regions of plasma cells from the brain of a patient with subacute sclerosing panencephalitis (SSPE). Five of eight human IgG1 recombinant antibodies (rAbs) derived from SSPE brain plasma cell clones recognized the measles virus (MV) nucleocapsid protein, confirming that the antibody response in SSPE targets primarily the agent causing disease. In this study, as part of our work on antigen identification, we used four rAbs to probe a random phage-displayed peptide library to determine if epitopes within the MV nucleocapsid protein could be identified with SSPE brain rAbs. All four of the SSPE rAbs enriched phage-displayed peptide sequences that reacted specifically to their panning rAb by enzyme-linked immunosorbent assay. BLASTP searches of the NCBI protein database revealed clear homologies in three peptides and different amino acid stretches within the 65 C-terminal amino acids of the MV nucleocapsid protein. The specificities of SSPE rAbs to these regions of the MV nucleocapsid protein were confirmed by binding to synthetic peptides or to short cDNA expression products. These results indicate the feasibility of using peptide screening for antigen discovery in central nervous system inflammatory diseases of unknown etiology, such as multiple sclerosis, neurosarcoidosis, or Behcet's syndrome.Panning of phage-displayed random peptide libraries allows an unbiased selection of antibody epitopes/mimotopes without preconceptions about the nature of the target antigens (reviewed in reference 23). In central nervous system (CNS) inflammatory diseases, where access to active diseased tissue is limited or where the levels of tissue antigen may be extremely low, phage peptide panning provides an alternative and sensitive avenue for antigen identification. Panning of phage-displayed random peptide libraries has successfully identified rheumatoid factor-specific mimotopes (22) and allergen mimotopes (19) and has mapped both linear and discontinuous viral epitopes recognized by antibodies specific for various infectious agents (5,10,11,16,20).Infectious and inflammatory diseases of the CNS are often characterized by increased intrathecal immunoglobulin G (IgG) synthesis that is seen as discrete bands of oligoclonal IgG when cerebrospinal fluid (CSF) or brain IgG is separated by isoelectric focusing. In CNS infectious diseases, such as subacute sclerosing panencephalitis (SSPE), neurosyphilis, mumps meningitis, progressive rubella panencephalitis, cryptococcal meningitis, and varicella zoster virus vasculitis, the oligoclonal IgG is directed largely against the infectious agent that causes the disease (reviewed in reference 12). Increased CNS IgG synthesis is accompanied by an elevated number of CD19 ϩ B cells and the app...

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Specificity of recombinant antibodies generated from multiple sclerosis cerebrospinal fluid probed with a random peptide library

Cited by 39 publications

References 39 publications

Clonally expanded plasma cells in the cerebrospinal fluid of MS patients produce myelin‐specific antibodies

Clonally expanded plasma cells in the cerebrospinal fluid of MS patients produce myelin‐specific antibodies

Antigen specificity of clonally expanded and receptor edited cerebrospinal fluid B cells from patients with relapsing remitting MS

Screening Random Peptide Libraries with Subacute Sclerosing PanencephalitisBrain-Derived Recombinant Antibodies Identifies Multiple Epitopes in the C-Terminal Region of the Measles Virus Nucleocapsid Protein

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