Epitope mapping and affinity purification of monospecific antibodies by Escherichia coli cell surface display of gene-derived random peptide libraries

Christmann, Andreas; Wentzel, Alexander; Meyer, Christiane; Meyers, Gregor; Kolmar, Harald

doi:10.1016/s0022-1759(01)00461-6

Cited by 51 publications

(37 citation statements)

References 28 publications

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“…that some residues are strongly conserved within the epitopes and suggested that these residues such as (11 aa) (Christmann et al, 2001;Zhang et al, 2006) and are distinct from the five groups of linear epitopes identified herein (Fig. 5a).…”

Section: Discussionmentioning

confidence: 55%

“…Five groups of CSFV E rns linear epitopes identified in this study are shown in underlined boldface and labelled I-V: group I for M2164, M2169, M2172 and M2178; group II for M2163, M2168 and M2171; group III for M2165 and M2177; group IV for M2167 and M2180; and group V for M2176. The overlapping epitopes recognized by mAb # 24/16 and mAb # b4-22 against the CSFV E rns (Christmann et al, 2001) are shown in boldface letters. The two regions of E rns constituting the active site of RNase are indicated by bars and annotated AS (aa 28-40) and AS (aa 71-89).…”

Section: Lin and Othersmentioning

confidence: 99%

“…Journal of General Virology 91 (Christmann et al, 2001) are present in close proximity within the same loop region. The group IV epitope recognized by M2167 and M2180 is located on helix 7, which is surface-exposed according to the 3D structural model of E rns .…”

Section: Lin and Others 2936mentioning

confidence: 99%

“…Eleven mAbs against CSFV E rns were established in previous studies (Kosmidou et al, 1995;Weiland et al, 1992). The epitopes for two of these 11 mAbs, mAb # b4-22 and mAb # 24/16, were mapped to two overlapping sequences 116 YDKNTDVNV 124 (9 aa) and 114 CRYDKNTDVNV 124 (11 aa) on the CSFV E rns , respectively, by using Escherichia coli cell surface display of gene-derived random peptide libraries (Christmann et al, 2001). Later, a study with the use of a phage display peptide library confirmed the epitopes and further revealed a binding motif 117 DKN 119 for both mAb # b4-22 and mAb # 24/16 (Zhang et al, 2006).…”

mentioning

confidence: 99%

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High-resolution epitope mapping for monoclonal antibodies to the structural protein Erns of classical swine fever virus using peptide array and random peptide phage display approaches

Lin

McRae

Dan

et al. 2010

Journal of General Virology

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The structural glycoprotein E rns (an envelope protein with RNase activity) of classical swine fever virus (CSFV) is not well characterized with respect to its antigenic structure and organization.Here, we investigated the antigenic sites on E rns by raising mAbs against the Escherichia coli expressed E rns of CSFV strain Alfort/187 and defined the B-cell epitopes recognized by these antibodies. Eighteen mAbs to E rns were identified and they were classified as either immunoglobulin subclass G1 or G2b. Using an array of overlapping 12-mer peptides, spanning aa 27-227 of E rns , the epitopes for 12 mAbs were mapped to a high resolution of six to eight residues, which cluster in five discrete locations, 31 GIWPEKIC 38 (group I), 65

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Section: Discussionmentioning

confidence: 55%

Section: Lin and Othersmentioning

confidence: 99%

Section: Lin and Others 2936mentioning

confidence: 99%

mentioning

confidence: 99%

See 2 more Smart Citations

High-resolution epitope mapping for monoclonal antibodies to the structural protein Erns of classical swine fever virus using peptide array and random peptide phage display approaches

Lin

McRae

Dan

et al. 2010

Journal of General Virology

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“…The supernatants of the hybridomas generated in this manner were screened for antigen binding and then tested for functionality in Western blots using RT-4 cell lysates and immunohistochemistry using tissue microarrays. 17 The Western blot analysis (Supporting Information Fig. 13) shows that monoclonal antibodies were obtained with the same staining as the corresponding monospecific antibodies.…”

Section: Generation Of Monoclonal Antibodies Against the Rbm3 Epitopementioning

confidence: 98%

Generation of monospecific antibodies based on affinity capture of polyclonal antibodies

et al. 2011

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A method is described to generate and validate antibodies based on mapping the linear epitopes of a polyclonal antibody followed by sequential epitope-specific capture using synthetic peptides. Polyclonal antibodies directed towards four proteins RBM3, SATB2, ANLN, and CNDP1, potentially involved in human cancers, were selected and antibodies to several non-overlapping epitopes were generated and subsequently validated by Western blot, immunohistochemistry, and immunofluorescence. For all four proteins, a dramatic difference in functionality could be observed for these monospecific antibodies directed to the different epitopes. In each case, at least one antibody was obtained with full functionality across all applications, while other epitope-specific fractions showed no or little functionality. These results present a path forward to use the mapped binding sites of polyclonal antibodies to generate epitope-specific antibodies, providing an attractive approach for large-scale efforts to characterize the human proteome by antibodies.

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Flow Cytometric Sorting of Bacterial Surface‐Displayed Libraries

2007

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The protocols herein detail methods for isolating binding peptides from a combinatorial library displayed on the surface of bacterial cells. These methods are appropriate for a variety of display scaffolds and a large range of library sizes, up to approximately 5 x 10(9) or more. Instructions have been provided for isolating peptides that bind to both proteins and non-protein targets, such as whole cells or inorganic particles. Qualitative analysis by flow cytometry can be exploited for bacterial libraries to characterize a displayed peptide's binding properties with a target of interest, and sorting conditions can be tuned to maximize binding affinity.

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Epitope mapping and affinity purification of monospecific antibodies by Escherichia coli cell surface display of gene-derived random peptide libraries

Cited by 51 publications

References 28 publications

High-resolution epitope mapping for monoclonal antibodies to the structural protein Erns of classical swine fever virus using peptide array and random peptide phage display approaches

High-resolution epitope mapping for monoclonal antibodies to the structural protein Erns of classical swine fever virus using peptide array and random peptide phage display approaches

Generation of monospecific antibodies based on affinity capture of polyclonal antibodies

Flow Cytometric Sorting of Bacterial Surface‐Displayed Libraries

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