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Keywords = MagPlex

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10 pages, 1773 KB  
Article
Selection and Characterization of Single-Domain Antibodies for Detection of Lassa Nucleoprotein
by George P. Anderson, Jinny L. Liu, Lisa C. Shriver-Lake and Ellen R. Goldman
Antibodies 2020, 9(4), 71; https://doi.org/10.3390/antib9040071 - 17 Dec 2020
Cited by 3 | Viewed by 4702
Abstract
Lassa virus is the etiologic agent of Lassa fever, an acute and often fatal illness endemic to West Africa. It is important to develop new reagents applicable either for the specific diagnosis or as improved therapeutics for the treatment of Lassa fever. Here, [...] Read more.
Lassa virus is the etiologic agent of Lassa fever, an acute and often fatal illness endemic to West Africa. It is important to develop new reagents applicable either for the specific diagnosis or as improved therapeutics for the treatment of Lassa fever. Here, we describe the development and initial testing of llama-derived single-domain antibodies that are specific for the Lassa virus nucleoprotein. Four sequence families based on complementarity-determining region (CDR) homology were identified by phage-based enzyme-linked immunosorbent assays, however, the highest affinity clones all belonged to the same sequence family which possess a second disulfide bond between Framework 2 and CDR3. The affinity and thermal stability were evaluated for each clone. A MagPlex-based homogeneous sandwich immunoassay for Lassa virus-like particles was also demonstrated to show their potential for further development as diagnostic reagents. Full article
(This article belongs to the Special Issue Design, Production and Characterization of Peptide Antibodies)
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12 pages, 3092 KB  
Article
Selection of Single-Domain Antibodies towards Western Equine Encephalitis Virus
by Jinny L. Liu, Lisa C. Shriver-Lake, Dan Zabetakis, Ellen R. Goldman and George P. Anderson
Antibodies 2018, 7(4), 44; https://doi.org/10.3390/antib7040044 - 15 Dec 2018
Cited by 15 | Viewed by 7278
Abstract
In this work, we describe the selection and characterization of single-domain antibodies (sdAb) towards the E2/E3E2 envelope protein of the Western equine encephalitis virus (WEEV). Our purpose was to identify novel recognition elements which could be used for the detection, diagnosis, and perhaps [...] Read more.
In this work, we describe the selection and characterization of single-domain antibodies (sdAb) towards the E2/E3E2 envelope protein of the Western equine encephalitis virus (WEEV). Our purpose was to identify novel recognition elements which could be used for the detection, diagnosis, and perhaps treatment of western equine encephalitis (WEE). To achieve this goal, we prepared an immune phage display library derived from the peripheral blood lymphocytes of a llama that had been immunized with an equine vaccine that includes killed WEEV (West Nile Innovator + VEWT). This library was panned against recombinant envelope (E2/E3E2) protein from WEEV, and seven representative sdAb from the five identified sequence families were characterized. The specificity, affinity, and melting point of each sdAb was determined, and their ability to detect the recombinant protein in a MagPlex sandwich immunoassay was confirmed. Thus, these new binders represent novel recognition elements for the E2/E3E2 proteins of WEEV that are available to the research community for further investigation into their applicability for use in the diagnosis or treatment of WEE. Full article
(This article belongs to the Special Issue Nanobody)
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15 pages, 1599 KB  
Article
Simultaneous Detection of Nine Key Bacterial Respiratory Pathogens Using Luminex xTAG® Technology
by Luxi Jiang, Hongyu Ren, Haijian Zhou, Tian Qin and Yu Chen
Int. J. Environ. Res. Public Health 2017, 14(3), 223; https://doi.org/10.3390/ijerph14030223 - 23 Feb 2017
Cited by 12 | Viewed by 5886
Abstract
Early diagnosis and treatment are crucial to the outcome of lower respiratory tract infections (LRTIs). In this study, we developed an assay combining multiplex PCR and Luminex technology (MPLT) for the detection of nine important respiratory bacterial pathogens, which frequently cause LRTIs. These [...] Read more.
Early diagnosis and treatment are crucial to the outcome of lower respiratory tract infections (LRTIs). In this study, we developed an assay combining multiplex PCR and Luminex technology (MPLT) for the detection of nine important respiratory bacterial pathogens, which frequently cause LRTIs. These were Streptococcus pneumoniae, Moraxella catarrhalis, Staphylococcus aureus, Streptococcus pyogenes, Haemophilus influenzae, Mycoplasma pneumoniae, Legionella spp., Pseudomonas aeruginosa, and Klebsiella pneumoniae. Through the hybridization reaction between two new synthesized multiplex PCR products and MagPlex-TAG Microspheres, we demonstrate that the detection limits for these nine pathogens were as low as 102–103 CFU/mL. Furthermore, 86 clinical bronchoalveolar lavage fluid specimens were used to evaluate this method. Compared with the results of nine simplex real-time PCR reactions targeting these nine pathogens, this MPLT assay demonstrated a high diagnostic accuracy for Streptococcus pneumoniae (sensitivity, 87.5% and specificity, 100%). Furthermore, sensitivity and specificity for the other eight pathogens all attained 100% diagnostic accuracy. In addition, the consistency between MPLT and the nine real-time PCR reactions exceeded 98.8%. In conclusion, MPLT is a high-throughput, labor-saving and reliable method with high sensitivity and specificity for identifying nine respiratory pathogens responsible for LRTIs. Indeed, this assay may be a promising supplement to conventional methods used to diagnose LRTIs. Full article
(This article belongs to the Section Environmental Health)
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