scholarly journals Iminodiacetic Acid (IDA) Cation-Exchange Nonwoven Membranes for Efficient Capture of Antibodies and Antibody Fragments

Membranes ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 530
Author(s):  
Jinxin Fan ◽  
Cristiana Boi ◽  
Solomon Mengistu Lemma ◽  
Joseph Lavoie ◽  
Ruben G. Carbonell
Keyword(s):  
Cation Exchange ◽  
Separation Efficiency ◽  
Antibody Fragment ◽  
Antibody Fragments ◽  
Residence Times ◽  
Single Chain ◽  
Dynamic Binding ◽  
Suitable Alternative ◽  
Wide Range ◽  
Weak Cation Exchange

There is strong need to reduce the manufacturing costs and increase the downstream purification efficiency of high-value therapeutic monoclonal antibodies (mAbs). This paper explores the performance of a weak cation-exchange membrane based on the coupling of IDA to poly(butylene terephthalate) (PBT) nonwoven fabrics. Uniform and conformal layers of poly(glycidyl methacrylate) (GMA) were first grafted to the surface of the nonwovens. Then IDA was coupled to the polyGMA layers under optimized conditions, resulting in membranes with very high permeability and binding capacity. This resulted in IgG dynamic binding capacities at very short residence times (0.1–2.0 min) that are much higher than those achieved by the best cation-exchange resins. Similar results were obtained in the purification of a single-chain (scFv) antibody fragment. As is customary with membrane systems, the dynamic binding capacities did not change significantly over a wide range of residence times. Finally, the excellent separation efficiency and potential reusability of the membrane were confirmed by five consecutive cycles of mAb capture from its cell culture harvest. The present work provides significant evidence that this weak cation-exchange nonwoven fabric platform might be a suitable alternative to packed resin chromatography for low-cost, higher productivity manufacturing of therapeutic mAbs and antibody fragments.

scholarly journals Optimization of the crystallizability of a single-chain antibody fragment

2014 ◽  
Vol 70 (12) ◽  
pp. 1701-1706 ◽  
Author(s):  
Jana Škerlová ◽  
Vlastimil Král ◽  
Milan Fábry ◽  
Juraj Sedláček ◽  
Václav Veverka ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Antibody Fragment ◽  
Binding Mode ◽  
Antibody Fragments ◽  
Dimensional Structure ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Diagnostic Potential ◽  
Storage Buffer ◽  
Single Chain Antibody Fragment

Single-chain variable antibody fragments (scFvs) are molecules with immense therapeutic and diagnostic potential. Knowledge of their three-dimensional structure is important for understanding their antigen-binding mode as well as for protein-engineering approaches such as antibody humanization. A major obstacle to the crystallization of single-chain variable antibody fragments is their relatively poor homogeneity caused by spontaneous oligomerization. A new approach to optimization of the crystallizability of single-chain variable antibody fragments is demonstrated using a representative single-chain variable fragment derived from the anti-CD3 antibody MEM-57. A Thermofluor-based assay was utilized to screen for optimal conditions for antibody-fragment stability and homogeneity. Such an optimization of the protein storage buffer led to a significantly improved ability of the scFv MEM-57 to yield crystals.

T-cell activation and cytokine production via a bispecific single-chain antibody fragment targeted to blood-stage malaria parasites

Blood ◽  
2003 ◽  
Vol 101 (6) ◽  
pp. 2300-2306 ◽  
Author(s):  
Shigeto Yoshida ◽  
Tominari Kobayashi ◽  
Hiroyuki Matsuoka ◽  
Chisato Seki ◽  
William L. Gosnell ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell Activation ◽  
Cell Activation ◽  
Recombinant Baculovirus ◽  
Antibody Fragment ◽  
Antibody Fragments ◽  
Therapeutic Drug ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Single Chain Antibody Fragment

A novel bispecific single-chain antibody fragment (biscFv) has been constructed to address the possibility of a new approach to malaria therapeutic drug development. The biscFv consists of 2 different single-chain antibody fragments linked by a flexible peptide linker (Gly4-Ser)3. Of the 2 scFv fragments, one is directed against a conserved epitope of the 19-kDa C-terminal fragment of the major surface protein of human malignant malaria parasite, Plasmodium falciparum, and the other is directed against the CD3 antigen of human T cells. The biscFv expressed by a recombinant baculovirus retained the antigen-binding properties of the corresponding univalent single-chain antibody fragments and formed a bridge between P falciparum and T cells. In cooperation with T cells, the biscFv specifically induced not only interferon γ and tumor necrosis factor α, but also a significant increase of merozoite phagocytosis and growth inhibition of P falciparum in vitro. Thus, the biscFv possesses highly selective malaria-targeting properties and stimulates T cells to induce cytokines, presumably resulting in activation of macrophages, neutrophils, and natural killer cells, and parasite killing in vivo.

scholarly journals Selection of Recombinant, Library-Derived Antibody Fragments against p24 for Human Immunodeficiency Virus Type 1 Diagnostics

1998 ◽  
Vol 5 (5) ◽  
pp. 636-644 ◽  
Author(s):  
Hans J. W. de Haard ◽  
Bert Kazemier ◽  
Marck J. M. Koolen ◽  
Liekle J. Nijholt ◽  
Rob H. Meloen ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Recombinant Antibody ◽  
Antibody Fragment ◽  
Antibody Fragments ◽  
Single Chain ◽  
Western Blots ◽  
Immunodeficiency Virus

ABSTRACT By application of combinatorial library technology, we generated the first recombinant antibody fragments directed against the major capsid protein p24 of human immunodeficiency virus type 1 (HIV-1). A library of single-chain Fv fragments (scFvs) was constructed by using the antibody variable-region (V) genes of B cells derived from the spleen of a viral lysate-immunized mouse. Antibodies were selected by panning or by enrichment with biotinylated antigen, yielding four different families of antibody fragments. The different types of scFvs were characterized by affinity measurements, by antigen recognition on Western blots, and by pepscan analysis. The epitope of one of the scFvs is located near the residues involved in CypA binding, thereby making it an attractive candidate for therapeutic applications. Comparison of the V gene sequence of this scFV with that of a previously described monoclonal antibody reactive against this immunodominant epitope revealed the usage of the identical combination of VH and Vκ regions. Thus, this is one of the rare examples in which the original combination in a library-derived antibody fragment was retrieved. After appropriate affinity and format improvements, the best of our recombinant scFvs may form the basis for a sensitive p24 assay as a measure of viral load. In addition, anti-p24 scFvs could be expressed as intracellular antibodies (intrabodies) to aid in the treatment of HIV infections.

scholarly journals Loxoscelism: Advances and Challenges in the Design of Antibody Fragments with Therapeutic Potential

Toxins ◽  
2020 ◽  
Vol 12 (4) ◽  
pp. 256
Author(s):  
Sabrina Karim-Silva ◽  
Alessandra Becker-Finco ◽  
Isabella Gizzi Jiacomini ◽  
Fanny Boursin ◽  
Arnaud Leroy ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Therapeutic Potential ◽  
Antibody Fragment ◽  
Binding Activity ◽  
Antibody Fragments ◽  
Antigen Binding ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Humanized Antibody ◽  
Neutralizing Capacity

Envenoming due to Loxosceles spider bites still remains a neglected disease of particular medical concern in the Americas. To date, there is no consensus for the treatment of envenomed patients, yet horse polyclonal antivenoms are usually infused to patients with identified severe medical conditions. It is widely known that venom proteins in the 30–35 kDa range with sphingomyelinase D (SMasesD) activity, reproduce most of the toxic effects observed in loxoscelism. Hence, we believe that monoclonal antibody fragments targeting such toxins might pose an alternative safe and effective treatment. In the present study, starting from the monoclonal antibody LimAb7, previously shown to target SMasesD from the venom of L. intermedia and neutralize its dermonecrotic activity, we designed humanized antibody V-domains, then produced and purified as recombinant single-chain antibody fragments (scFvs). These molecules were characterized in terms of humanness, structural stability, antigen-binding activity, and venom-neutralizing potential. Throughout this process, we identified some blocking points that can impact the Abs antigen-binding activity and neutralizing capacity. In silico analysis of the antigen/antibody amino acid interactions also contributed to a better understanding of the antibody’s neutralization mechanism and led to reformatting the humanized antibody fragment which, ultimately, recovered the functional characteristics for efficient in vitro venom neutralization.

scholarly journals A microbial expression system for high-level production of scFv HIV-neutralizing antibody fragments in Escherichia coli

2019 ◽  
Vol 103 (21-22) ◽  
pp. 8875-8888 ◽  
Author(s):  
Marloes L. C. Petrus ◽  
Lukas A. Kiefer ◽  
Pranav Puri ◽  
Evert Heemskerk ◽  
Michael S. Seaman ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Neutralizing Antibodies ◽  
Expression System ◽  
Antibody Fragment ◽  
Green Fluorescence Protein ◽  
Antibody Fragments ◽  
Single Chain ◽  
Mammalian Expression ◽  
Anti Hiv

Abstract Monoclonal antibodies (mABs) are of great biopharmaceutical importance for the diagnosis and treatment of diseases. However, their production in mammalian expression hosts usually requires extensive production times and is expensive. Escherichia coli has become a new platform for production of functional small antibody fragment variants. In this study, we have used a rhamnose-inducible expression system that allows precise control of protein expression levels. The system was first evaluated for the cytoplasmic production of super folder green fluorescence protein (sfGFP) in various production platforms and then for the periplasmic production of the anti-HIV single-chain variable antibody fragment (scFv) of PGT135. Anti-HIV broadly neutralizing antibodies, like PGT135, have potential for clinical use to prevent HIV transmission, to promote immune responses and to eradicate infected cells. Different concentrations of L-rhamnose resulted in the controlled production of both sfGFP and scFv PGT135 antibody. In addition, by optimizing the culture conditions, the amount of scFv PGT135 antibody that was expressed soluble or as inclusions bodies could be modulated. The proteins were produced in batch bioreactors, with yields of 4.9 g/L for sfGFP and 0.8 g/L for scFv. The functionality of the purified antibodies was demonstrated by their ability to neutralize a panel of different HIV variants in vitro. We expect that this expression system will prove very useful for the development of a more cost-effective production process for proteins and antibody fragments in microbial cells.

Antibody fragment and targeted colorectal cancer therapy: A global systematic review

2021 ◽  
Vol 22 ◽  
Author(s):  
Sepideh Ghani ◽  
Niloofar Deravi ◽  
Marzieh Pirzadeh ◽  
Behnam Rafiee ◽  
Zahra Rezanejad Gatabi ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Systematic Review ◽  
Immune System ◽  
Tumor Growth ◽  
Cancer Treatment ◽  
Antibody Fragment ◽  
Antibody Fragments ◽  
Great Promise ◽  
Single Chain

Background and aims: Antibody-based therapeutics have been evidenced promising for the treatment of colorectal cancer patients. However, the size and long circulating half-lives of antibodies can limit their reproducible manufacture in clinical studies. Consequently, in novel therapeutic approaches conventional antibodies are minimized and engineered to produce fragments like Fab, scFv, nanobody, bifunctional antibody, bispecific antibody, minibody and diabody to preserve their high affinity and specificity to target pharmaceutical nanoparticle conjugates. This systematic review for the first time aimed to elucidate the role of various antibody fragments in colorectal cancer treatment. Method: A systematic literature search in web of sciences, PubMed, Scopus, Google scholar and ProQuest was conducted. Reference lists of the articles were reviewed to identify the relevant papers. The full text search included articles published in English during 1990–2021. Results: Most the 53 included studies were conducted in vitro and in most conducted studies single-chain antibodies were among the most used antibody fragments. Most antibodies targeted CEA in the treatment of colorectal cancer. Moreover, a large number of studies observed apoptosis induction and tumor growth inhibition. In addition, few studies implicated the role of the innate immune system as an indirect mechanisms of tumor growth by enhancing NK-cell killing. Conclusion: Antibody-based therapy was demonstrated to be of a great promise in the treatment of colorectal cancer rather than common treatments such as radiotherapy, chemotherapy, and surgical operations. This type of specified cancer treatment can also induce the activation of innate and specific immune system to eradicate tumor cells.

scholarly journals Single Chain Variable Fragment Fused to Maltose Binding Protein: A Modular Nanocarrier Platform for the Targeted Delivery of Antitumorals

2021 ◽  
Author(s):  
Francisco J. Reche-Perez ◽  
Simona Plesselova ◽  
Eduardo De los Reyes-Berbel ◽  
Mariano Ortega-Muñoz ◽  
F. Javier Lopez-Jaramillo ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Targeted Delivery ◽  
Specific Binding ◽  
Protein A ◽  
Antibody Fragments ◽  
Single Chain Variable Fragment ◽  
Single Chain ◽  
Binding Properties ◽  
Single Chain Variable Fragments ◽  
Selective Delivery

The use of the specific binding properties of monoclonal antibody fragments such as single-chain variable fragments (ScFv) for the selective delivery of antitumor therapeutics for cancer cells is attractive due...

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