scholarly journals Genetic Fusion of an Anti-BclA Single-Domain Antibody with Beta Galactosidase

Antibodies ◽  
2018 ◽  
Vol 7 (4) ◽  
pp. 36 ◽  
Author(s):  
George Anderson ◽  
Lisa Shriver-Lake ◽  
Scott Walper ◽  
Lauryn Ashford ◽  
Dan Zabetakis ◽  
...  
Keyword(s):  
Single Domain ◽  
Single Domain Antibody ◽  
Domain Antibody ◽  
Highly Sensitive ◽  
Bacillus Anthracis Spores ◽  
Anthrax Spores ◽  
Single Domain Antibodies ◽  
Genetic Fusion ◽  
Sandwich Assays ◽  
Beta Galactosidase

The Bacillus collagen-like protein of anthracis (BclA), found in Bacillus anthracis spores, is an attractive target for immunoassays. Previously, using phage display we had selected llama-derived single-domain antibodies that bound to B. anthracis spore proteins including BclA. Single-domain antibodies (sdAbs), the recombinantly expressed heavy domains from the unique heavy-chain-only antibodies found in camelids, provide stable and well-expressed binding elements with excellent affinity. In addition, sdAbs offer the important advantage that they can be tailored for specific applications through protein engineering. A fusion of a BclA targeting sdAb with the enzyme Beta galactosidase (β-gal) would enable highly sensitive immunoassays with no need for a secondary reagent. First, we evaluated five anti-BclA sdAbs, including four that had been previously identified but not characterized. Each was tested to determine its binding affinity, melting temperature, producibility, and ability to function as both capture and reporter in sandwich assays for BclA. The sdAb with the best combination of properties was constructed as a fusion with β-gal and shown to enable sensitive detection. This fusion has the potential to be incorporated into highly sensitive assays for the detection of anthrax spores.

scholarly journals Retargeting of Adenovirus Vectors through Genetic Fusion of a Single-Chain or Single-Domain Antibody to Capsid Protein IX

2010 ◽  
Vol 84 (19) ◽  
pp. 10074-10086 ◽  
Author(s):  
Kathy L. Poulin ◽  
Robert M. Lanthier ◽  
Adam C. Smith ◽  
Carin Christou ◽  
Milagros Risco Quiroz ◽  
...  
Keyword(s):  
Gene Therapy ◽  
Virus Infection ◽  
Capsid Protein ◽  
Cell Types ◽  
Single Domain ◽  
Single Chain ◽  
Single Domain Antibody ◽  
Single Domain Antibodies ◽  
Genetic Fusion ◽  
Protein Ix

ABSTRACT Adenovirus (Ad) vectors are the most commonly used system for gene therapy applications, due in part to their ability to infect a wide array of cell types and tissues. However, many therapies would benefit from the ability to target the Ad vector only to specific cells, such as tumor cells for cancer gene therapy. In this study, we investigated the utility of capsid protein IX (pIX) as a platform for the presentation of single-chain variable-fragment antibodies (scFv) and single-domain antibodies (sdAb) for virus retargeting. We show that scFv can be displayed on the capsid through genetic fusion to native pIX but that these molecules fail to retarget the virus, due to improper folding of the scFv. Redirecting expression of the fusion protein to the endoplasmic reticulum (ER) results in correct folding of the scFv and allows it to recognize its epitope; however, ER-targeted pIX-scFv was incorporated into the Ad capsid at a very low level which was not sufficient to retarget virus infection. In contrast, a pIX-sdAb construct was efficiently incorporated into the Ad capsid and enhanced virus infection of cells expressing the targeted receptor. Taken together, our data indicate that pIX is an effective platform for presentation of large targeting polypeptides on the surface of the virus capsid, but the nature of the ligand can significantly affect its association with virions.

scholarly journals Single-Domain Antibody-Based TCR-Like CAR-T: A Potential Cancer Therapy

2020 ◽  
Vol 2020 ◽  
pp. 1-8
Author(s):  
Lichen Zhu ◽  
Xiaomei Yang ◽  
Dani Zhong ◽  
Shenxia Xie ◽  
Wei Shi ◽  
...  
Keyword(s):  
T Cells ◽  
Hematological Malignancies ◽  
Single Domain ◽  
Current Status ◽  
Great Success ◽  
Single Domain Antibody ◽  
Domain Antibody ◽  
Car T ◽  
Single Domain Antibodies ◽  
Potential Cancer

Retargeting the antigen-binding specificity of T cells to intracellular antigens that are degraded and presented on the tumor surface by engineering chimeric antigen receptor (CAR), also named TCR-like antibody CAR-T, remains limited. With the exception of the commercialized CD19 CAR-T for hematological malignancies and other CAR-T therapies aiming mostly at extracellular antigens achieving great success, the rareness and scarcity of TCR-like CAR-T therapies might be due to their current status and limitations. This review provides the probable optimized initiatives for improving TCR-like CAR-T reprogramming and discusses single-domain antibodies administered as an alternative to conventional scFvs and secreted by CAR-T cells, which might be of great value to the development of CAR-T immunotherapies for intracellular antigens.

scholarly journals Rugged Single Domain Antibody Detection Elements for Bacillus anthracis Spores and Vegetative Cells

PLoS ONE ◽  
2012 ◽  
Vol 7 (3) ◽  
pp. e32801 ◽  
Author(s):  
Scott A. Walper ◽  
George P. Anderson ◽  
P. Audrey Brozozog Lee ◽  
Richard H. Glaven ◽  
Jinny L. Liu ◽  
...  
Keyword(s):  
Bacillus Anthracis ◽  
Single Domain ◽  
Antibody Detection ◽  
Single Domain Antibody ◽  
Vegetative Cells ◽  
Domain Antibody ◽  
Bacillus Anthracis Spores

scholarly journals Fully human single-domain antibodies against SARS-CoV-2

2020 ◽  
Author(s):  
Yanling Wu ◽  
Cheng Li ◽  
Shuai Xia ◽  
Xiaolong Tian ◽  
Zhi Wang ◽  
...  
Keyword(s):  
Single Domain ◽  
Human Origin ◽  
Antibody Library ◽  
Single Domain Antibody ◽  
Live Virus ◽  
Efficient Approach ◽  
High Affinity ◽  
Domain Antibody ◽  
Single Domain Antibodies ◽  
Neutralizing Epitopes

AbstractThe COVID-19 pandemic is spreading rapidly, highlighting the urgent need for an efficient approach to rapidly develop therapeutics and prophylactics against SARS-CoV-2. We describe here the development of a phage-displayed single-domain antibody library by grafting naïve CDRs into framework regions of an identified human germline IGHV allele. This enabled the isolation of high-affinity single-domain antibodies of fully human origin. The panning using SARS-CoV-2 RBD and S1 as antigens resulted in the identification of antibodies targeting five types of neutralizing or non-neutralizing epitopes on SARS-CoV-2 RBD. These fully human single-domain antibodies bound specifically to SARS-CoV-2 RBD with subnanomolar to low nanomolar affinities. Some of them were found to potently neutralize pseudotyped and live virus, and therefore may represent promising candidates for prophylaxis and therapy of COVID-19. This study also reports unique immunogenic profile of SARS-CoV-2 RBD compared to that of SARS-CoV and MERS-CoV, which may have important implications for the development of effective vaccines against SARS-CoV-2.

Highly Sensitive Single Domain Antibody–Quantum Dot Conjugates for Detection of HER2 Biomarker in Lung and Breast Cancer Cells

ACS Nano ◽  
2014 ◽  
Vol 8 (6) ◽  
pp. 5682-5695 ◽  
Author(s):  
Tatsiana Y. Rakovich ◽  
Omar K. Mahfoud ◽  
Bashir M. Mohamed ◽  
Adriele Prina-Mello ◽  
Kieran Crosbie-Staunton ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Quantum Dot ◽  
Cancer Cells ◽  
Breast Cancer Cells ◽  
Single Domain ◽  
Single Domain Antibody ◽  
Domain Antibody ◽  
Highly Sensitive

Production, characterization and in-vitro applications of single-domain antibody against thyroglobulin selected from novel T7 phage display library

2021 ◽  
Vol 492 ◽  
pp. 112990
Author(s):  
Jothivel Kumarasamy ◽  
Samar Kumar Ghorui ◽  
Chandrakala Gholve ◽  
Bharti Jain ◽  
Yogesh Dhekale ◽  
...  
Keyword(s):  
Phage Display ◽  
Phage Display Library ◽  
Single Domain ◽  
Single Domain Antibody ◽  
Domain Antibody ◽  
T7 Phage Display ◽  
T7 Phage

scholarly journals A Single-Domain Antibody-Based Anti-PSMA Recombinant Immunotoxin Exhibits Specificity and Efficacy for Prostate Cancer Therapy

2021 ◽  
Vol 22 (11) ◽  
pp. 5501
Author(s):  
Yutong Xing ◽  
Keyuan Xu ◽  
Shixiong Li ◽  
Li Cao ◽  
Yue Nan ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Animal Studies ◽  
Single Domain ◽  
Soluble Form ◽  
Therapeutic Window ◽  
Single Domain Antibody ◽  
Pseudomonas Exotoxin A ◽  
Simple Strategy ◽  
Domain Antibody ◽  
Recombinant Immunotoxin

Prostate cancer (PCa) is the second most common cancer in men, causing more than 300,000 deaths every year worldwide. Due to their superior cell-killing ability and the relative simplicity of their preparation, immunotoxin molecules have great potential in the clinical treatment of cancer, and several such molecules have been approved for clinical application. In this study, we adopted a relatively simple strategy based on a single-domain antibody (sdAb) and an improved Pseudomonas exotoxin A (PE) toxin (PE24X7) to prepare a safer immunotoxin against prostate-specific membrane antigen (PSMA) for PCa treatment. The designed anti-PSMA immunotoxin, JVM-PE24X7, was conveniently prepared in its soluble form in an Escherichia coli (E. coli) system, avoiding the complex renaturation process needed for immunotoxin preparation by the conventional strategy. The product was very stable and showed a very strong ability to bind the PSMA receptor. Cytotoxicity assays showed that this molecule at a very low concentration could kill PSMA-positive PCa cells, with an EC50 value (concentration at which the cell viability decreased by 50%) of 15.3 pM against PSMA-positive LNCaP cells. Moreover, this molecule showed very good killing selectivity between PSMA-positive and PSMA-negative cells, with a selection ratio of more than 300-fold. Animal studies showed that this molecule at a very low dosage (5 × 0.5 mg/kg once every three days) completely inhibited the growth of PCa tumors, and the maximum tolerable dose (MTD) was more than 15 mg/kg, indicating its very potent tumor-treatment ability and a wide therapeutic window. Use of the new PE toxin, PE24X7, as the effector moiety significantly reduced off-target toxicity and improved the therapeutic window of the immunotoxin. The above results demonstrate that the designed anti-PSMA immunotoxin, JVM-PE24X7, has good application value for the treatment of PCa.

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