antibody phage display

Monoclonal antibodies (mAbs) are one of the most important classes of biologics with high therapeutic and diagnostic value, but traditional methods for mAbs generation, such as hybridoma screening and phage display, have limitations, including low efficiency and loss of natural chain pairing. To overcome these challenges, novel single B cell antibody technologies have emerged, but they also have limitations such as in vitro differentiation of memory B cells and expensive cell sorters. In this study, we present a rapid and efficient workflow for obtaining human recombinant monoclonal antibodies directly from single antigen-specific antibody secreting cells (ASCs) in the peripheral blood of convalescent COVID-19 patients using ferrofluid technology...

The emergence of the Middle East Respiratory Syndrome Coronavirus (MERS-CoV) has posed a significant global health concern due to its severe respiratory illness and high fatality rate. Currently, despite the potential for resurgence, there are no specific treatments for MERS-CoV, and only supportive care is available. Our study aimed to address this therapeutic gap by developing a potent neutralizing bispecific antibody (bsAb) against MERS-CoV. Initially, we isolated four human monoclonal antibodies (mAbs) that specifically target the MERS-CoV receptor-binding domain (RBD) using phage display technology and an established human antibody library...

Immunogenicity is a major caveat of protein therapeutics. In particular, the long-term administration of protein therapeutic agents leads to the generation of antidrug antibodies (ADAs), which reduce drug efficacy while eliciting adverse events. One promising solution to this issue is the use of mirror-image proteins consisting of d-amino acids, which are resistant to proteolytic degradation in immune cells. We have recently reported the chemical synthesis of the enantiomeric form of the variable domain of the antibody heavy chain (d-VHH)...

SARS-CoV-2 continues to threaten human health, antibody therapy is one way to control the infection. Because new SARS-CoV-2 mutations are constantly emerging, there is an urgent need to develop broadly neutralizing antibodies to block the viral entry into host cells. VNAR from sharks is the smallest natural antigen binding domain, with the advantages of small size, flexible paratopes, good stability, and low manufacturing cost. Here, we used recombinant SARS-CoV-2 Spike-RBD to immunize sharks and constructed a VNAR phage display library...

Human immunoglobulin preparations contain a diverse range of polyclonal antibodies that reflect past immune responses against pathogens encountered by the blood donor population. In this study, we examined a panel of intravenous immunoglobulins (IGIVs) manufactured over the past two decades (1998-2020) for their capacity to neutralize or enhance Zika virus (ZIKV) infection in vitro . These IGIVs were selected specifically based on their production dates in relation to the occurrences of two flavivirus outbreaks in the U...

Directing antibodies to a particular epitope among many possible on a target protein is a significant challenge. Here, we present a simple and general method for epitope-directed selection (EDS) using a differential phage selection strategy. This involves engineering the protein of interest (POI) with the epitope of interest (EOI) mutated using a systematic bioinformatics algorithm to guide the local design of an EOI decoy variant. Using several alternating rounds of negative selection with the EOI decoy variant followed by positive selection on the wild-type POI, we were able to identify highly specific and potent antibodies to five different EOI antigens that bind and functionally block known sites of proteolysis...

Phage display has become an efficient, reliable and popular molecular technique for generating libraries encompassing millions or even billions of clones of divergent peptides or proteins. The method is based on the correspondence between phage genotype and phenotype, which ensures the presentation of recombinant proteins of known amino acid composition on the surface of phage particles. The use of affinity selection allows one to choose variants with affinity for different targets from phage libraries. The implementation of the antibody phage display technique has revolutionized the field of clinical immunology, both for developing tools to diagnose infectious diseases and for producing therapeutic agents...

Antibodies that specifically bind to individual human fragment crystallizable γ receptors (FcγRs) are of interest as research tools in studying immune cell functions, as well as components in bispecific antibodies for immune cell engagement in cancer therapy. Monoclonal antibodies for human low-affinity FcγRs have been successfully generated by hybridoma technology and are widely used in pre-clinical research. However, the generation of monoclonal antibodies by hybridoma technology that specifically bind to the high-affinity receptor FcγRI is challenging...

Most available neutralizing antibodies are ineffective against highly mutated SARS-CoV-2 Omicron subvariants. Therefore, it is crucial to develop potent and broad-spectrum alternatives to effectively manage Omicron subvariants. Here, we constructed a high-diversity nanobody phage display library and identified nine nanobodies specific to the SARS-CoV-2 receptor-binding domain (RBD). Five of them exhibited cross-neutralization activity against the SARS-CoV-2 wild-type (WT) strain and the Omicron subvariants BA...

The variable domain of a heavy-chain antibody (VHH) has the potential to be used to redirect the cell tropism of adenoviral vectors. Here, we attempted to establish platforms to simplify the screening of VHHs for their specific targeting function when being incorporated into the fiber of adenovirus. Both fowl adenovirus 4 (FAdV-4) and simian adenovirus 1 (SAdV-1) have two types of fiber, one of which is dispensable for virus propagation and is a proper site for VHH display. An intermediate plasmid, pMD-FAV4Fs, was constructed as the start plasmid for FAdV-4 fiber2 modification...

Nanobodies have emerged as promising tools in biomedicine due to their single-chain structure and inherent stability. They generally have convex paratopes, which potentially prefer different epitope sites in an antigen compared to traditional antibodies. In this study, a synthetic phage display nanobody library was constructed and used to identify nanobodies targeting a tumor-associated antigen, the human B7-H3 protein. Combining next-generation sequencing and single-clone validation, two nanobodies were identified to specifically bind B7-H3 with medium nanomolar affinities...

When purifying mAb from serum-containing hybridoma culture supernatant, it is essential that mouse IgG remains free from contaminations of bovine IgG. However, the broadly used Protein A resin cannot achieve this goal due to binding between both mouse and bovine IgG. Here, a novel nanobody-based affinity purification magnetic beads that discriminates mouse IgG from bovine IgG was developed. To bind all subtypes of mouse IgG (IgG1, IgG2a, IgG2b and IgG3) that contain the kappa light chain, mCK (mouse kappa constant region)-specific nanobody binders were selected from an immune phage display VHH library; this library was constructed with peripheral blood mononuclear cells (PBMCs), which were collected from Bactrian camels immunized with a mix of intact mouse IgGs (IgG1, IgG2a, IgG2b and IgG3)...

Cancer immunotherapy targeting adaptive immune cells has been attracting considerable interest due to its great success in treating multiple cancers. Recently, there is also increasing interest in agents that can stimulate innate immune cell activities. Immune checkpoint inhibitors targeting innate immune cells can block inhibitory interactions ('don't eat me' signals) between tumor cells and phagocytes. CD47 is a transmembrane protein overexpressed in various cancers and acts as a potent 'do not eat me' signal that contributes to the immune evasion of cancer cells...

BACKGROUND: Rational design of synthetic phage-displayed libraries requires the identification of the most appropriate positions for randomization using defined amino acid sets to recapitulate the natural occurrence. The present study uses position-specific scoring matrixes (PSSMs) for identifying and randomizing Camelidae nanobody (VHH) CDR3. The functionality of a synthetic VHH repertoire designed by this method was tested for discovering new VHH binders to recombinant coagulation factor VII (rfVII)...

There is no doubt that today's life sciences would look very different without the availability of millions of research antibody products. Nevertheless, the use of antibody reagents that are poorly characterized has led to the publication of false or misleading results. The use of laboratory animals to produce research antibodies has also been criticized. Surprisingly, both problems can be addressed with the same technology. This review charts today's maze of different antibody formats and the various methods for antibody production and their interconnections, ultimately concluding that sequence-defined recombinant antibodies offer a clear path to both improved quality of experimental data and reduced use of animals...

Phage display technology has become an important research tool in biological research, fundamentally changing the traditional monoclonal antibody preparation process, and has been widely used in the establishment of antigen-antibody libraries, drug design, vaccine research, pathogen detection, gene therapy, antigenic epitope research, and cellular signal transduction research.The phage display is a powerful platform for technology development. Using phage display technology, single chain fragment variable (scFv) can be screened, replacing the disadvantage of the large size of traditional antibodies...

Protein phosphorylation is a prevalent translational modification, and its dysregulation has been implicated in various diseases, including cancer. Despite its significance, there is a lack of specific inhibitors of the FCP/SCP-type Ser/Thr protein phosphatase Scp1, characterized by high specificity and affinity. In this study, we focused on adnectin, an antibody-mimetic protein, aiming to identify Scp1-specific binding molecules with a broad binding surface that target the substrate-recognition site of Scp1...

The injudicious usage of antibiotics during infections caused by Gram-negative bacteria leads to the emergence of β-lactamases. Among them, the NDM-1 enzyme poses a serious threat to human health. Developing new antibiotics or inhibiting β-lactamases might become essential to reduce and prevent bacterial infections. Nanobodies (Nbs), the smallest antigen-binding single-domain fragments derived from Camelidae heavy-chain-only antibodies, targeting enzymes, are innovative alternatives to develop effective inhibitors...

Advanced biopharmaceutical manufacturing requires novel process analytical technologies for the rapid and sensitive assessment of the higher-order structures of therapeutic proteins. However, conventional physicochemical analyses of denatured proteins have limitations in terms of sensitivity, throughput, analytical resolution, and real-time monitoring capacity. Although probe-based sensing can overcome these limitations, typical non-specific probes lack analytical resolution and provide little to no information regarding which parts of the protein structure have been collapsed...

BACKGROUND: Diphtheria can be prevented by vaccination, but some epidemics occur in several places, and diphtheria's threat is considerable. Administration of diphtheria antitoxin (DAT) produced from hyperimmunized animals is the most common treatment. Recombinant human antibody fragments such as single-chain variable fragments (scFv) produced by phage display library may introduce an interesting approach to overcome the limitations of the traditional antibody therapy. In the present study, B cells of immunized volunteers were used to construct a human single-chain fragment (HuscFv) library...