Assessing Antibody Specificity in Human Serum Using Deep Sequence-Coupled Biopanning

Frietze, Kathryn M.; Core, Susan B.; Linville, Alexandria C; Chackerian, Bryce; Peabody, David S.

doi:10.1007/978-1-4939-9853-1_9

Cited by 7 publications

(10 citation statements)

References 9 publications

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“…After the fourth round of selection, individual transformants were isolated and sequenced. A more detailed protocol describing the generation of VLPs displaying random peptides and affinity selections is also available ( 60 ).…”

Section: Methodsmentioning

confidence: 99%

Engineering an Antibody V Gene-Selective Vaccine

et al. 2021

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The ligand-binding surface of the B cell receptor (BCR) is formed by encoded and non-encoded antigen complementarity determining regions (CDRs). Genetically reproducible or ‘public’ antibodies can arise when the encoded CDRs play deterministic roles in antigen recognition, notably within human broadly neutralizing antibodies against HIV and influenza virus. We sought to exploit this by engineering virus-like-particle (VLP) vaccines that harbor multivalent affinity against gene-encoded moieties of the BCR antigen binding site. As proof of concept, we deployed a library of RNA bacteriophage VLPs displaying random peptides to identify a multivalent antigen that selectively triggered germline BCRs using the human VH gene IGVH1-2*02. This VLP selectively primed IGHV1-2*02 BCRs that were present within a highly diversified germline antibody repertoire within humanized mice. Our approach thus provides methodology to generate antigens that engage specific BCR configurations of interest, in the absence of structure-based information.

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Section: Methodsmentioning

confidence: 99%

Engineering an Antibody V Gene-Selective Vaccine

et al. 2021

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“…The method used for the deep sequence-coupled biopanning has been previously described [ 1 , 2 , 27 ] and had the following modifications. IgG was isolated from 20 µL of patient sera using Dynabeads Protein G (Invitrogen) following the manufacturer’s instructions.…”

Section: Methodsmentioning

confidence: 99%

“…The purified PCR product was then subjected to Ion Torrent deep sequencing. Raw data were processed with custom MATLAB scripts as previously described [ 1 , 2 , 27 ]. Sequences that passed quality control standards were then used to identify sequences encoding unique peptides and rank them according to their abundance (% total population of quality-controlled sequences).…”

Section: Methodsmentioning

confidence: 99%

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Expansion and Refinement of Deep Sequence-Coupled Biopanning Technology for Epitope-Specific Antibody Responses in Human Serum

Warner

Linville

Core

et al. 2020

Viruses

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Identifying the specific epitopes targeted by antibodies elicited in response to infectious diseases is important for developing vaccines and diagnostics. However, techniques for broadly exploring the specificity of antibodies in a rapid manner are lacking, limiting our ability to quickly respond to emerging viruses. We previously reported a technology that couples deep sequencing technology with a bacteriophage MS2 virus-like particle (VLP) peptide display platform for identifying pathogen-specific antibody responses. Here, we describe refinements that expand the number of patient samples that can be processed at one time, increasing the utility of this technology for rapidly responding to emerging infectious diseases. We used dengue virus (DENV) as a model system since much is already known about the antibody response. Sera from primary DENV-infected patients (n = 28) were used to pan an MS2 bacteriophage VLP library displaying all possible 10-amino-acid peptides from the DENV polypeptide. Selected VLPs were identified by deep sequencing and further investigated by enzyme-linked immunosorbent assay. We identified previously described immunodominant regions of envelope and nonstructural protein-1, as well as a number of other epitopes. Our refinement of the deep sequence-coupled biopanning technology expands the utility of this approach for rapidly investigating the specificity of antibody responses to infectious diseases.

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“…MS2 VLPs, in particular, are amendable to the display of short peptide epitopes in a constrained surface-exposed loop, mimicking conformational epitopes that naturally form loop structures [21,24,27]. Furthermore, MS2 VLPs can be used in a novel affinity selection platform called Deep Sequence-Coupled Biopanning [23,[28][29][30][31]. Using this technology, we can identify conformational mimics that bind to a specific monoclonal antibody [23].…”

Section: Introductionmentioning

confidence: 99%

Epitope-Based Vaccines against the Chlamydia trachomatis Major Outer Membrane Protein Variable Domain 4 Elicit Protection in Mice

et al. 2022

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Chlamydia trachomatis (Ct) is the most common bacterial sexual transmitted pathogen, yet a vaccine is not currently available. Here, we used the immunogenic bacteriophage MS2 virus-like particle (VLP) technology to engineer vaccines against the Ct major outer membrane protein variable domain 4 (MOMP-VD4), which contains a conserved neutralizing epitope (TTLNPTIAG). A previously described monoclonal antibody to the MOMP-VD4 (E4 mAb) is capable of neutralizing all urogenital Ct serovars and binds this core epitope, as well as several non-contiguous amino acids. This suggests that this core epitope may require conformational context in order to elicit neutralizing antibodies to Ct. In order to identify immunogens that could elicit neutralizing antibodies to the TTLNPTIAG epitope, we used two approaches. First, we used affinity selection with a bacteriophage MS2-VLP library displaying random peptides in a constrained, surface-exposed loop to identify potential E4 mAb mimotopes. After four rounds of affinity selection, we identified a VLP-displayed peptide (HMVGSTKWTN) that could bind to the E4 mAb and elicited serum IgG that bound weakly to Ct elementary bodies by ELISA. Second, two versions of the core conserved TTLNPTIAG epitope (TTLNPTIAG and TTLNPTIAGA) were recombinantly expressed on the coat protein of the MS2 VLP in a constrained, surface-exposed loop. Mouse immune sera IgG bound to Ct elementary bodies by ELISA. Immunization with these MS2 VLPs provided protection from vaginal Chlamydia infection in a murine challenge model. These data suggest that short peptide epitopes targeting the MOMP-VD4 could be appropriate for Ct vaccine design when displayed on an immunogenic bacteriophage VLP vaccine platform.

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Assessing Antibody Specificity in Human Serum Using Deep Sequence-Coupled Biopanning

Cited by 7 publications

References 9 publications

Engineering an Antibody V Gene-Selective Vaccine

Engineering an Antibody V Gene-Selective Vaccine

Expansion and Refinement of Deep Sequence-Coupled Biopanning Technology for Epitope-Specific Antibody Responses in Human Serum

Epitope-Based Vaccines against the Chlamydia trachomatis Major Outer Membrane Protein Variable Domain 4 Elicit Protection in Mice

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