Biolayer Interferometry: Protein–RNA Interactions

2021 ◽  
pp. 351-368
Author(s):  
Stephen R. Martin ◽  
Andres Ramos ◽  
Laura Masino
Keyword(s):  
Biolayer Interferometry

scholarly journals Homogenous overexpression of the extracellular matrix protein Netrin-1 in a hollow fiber bioreactor

2021 ◽  
Author(s):  
Aniel Moya-Torres ◽  
Monika Gupta ◽  
Fabian Heide ◽  
Natalie Krahn ◽  
Scott Legare ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Hollow Fiber ◽  
Mammalian Cells ◽  
Matrix Protein ◽  
Continuous Production ◽  
Extracellular Matrix Protein ◽  
Close Monitoring ◽  
High Quality ◽  
Biolayer Interferometry ◽  
Hollow Fiber Bioreactor

Abstract The production of recombinant proteins for functional and biophysical studies, especially in the field of structural determination, still represents a challenge as high quality and quantities are needed to adequately perform experiments. This is in part solved by optimizing protein constructs and expression conditions to maximize the yields in regular flask expression systems. Still, work flow and effort can be substantial with no guarantee to obtain improvements. This study presents a combination of workflows that can be used to dramatically increase protein production and improve processing results, specifically for the extracellular matrix protein Netrin-1. This proteoglycan is an axon guidance cue which interacts with various receptors to initiate downstream signaling cascades affecting cell differentiation, proliferation, metabolism, and survival. We were able to produce large glycoprotein quantities in mammalian cells, which were engineered for protein overexpression and secretion into the media using the controlled environment provided by a hollow fiber bioreactor. Close monitoring of the internal bioreactor conditions allowed for stable production over an extended period of time. In addition to this, Netrin-1 concentrations were monitored in expression media through biolayer interferometry which allowed us to increase Netrin-1 media concentrations tenfold over our current flask systems while preserving excellent protein quality and in solution behavior. Our particular combination of genetic engineering, cell culture system, protein purification, and biophysical characterization permitted us to establish an efficient and continuous production of high-quality protein suitable for structural biology studies that can be translated to various biological systems. Key points • Hollow fiber bioreactor produces substantial yields of homogenous Netrin-1 • Biolayer interferometry allows target protein quantitation in expression media • High production yields in the bioreactor do not impair Netrin-1 proteoglycan quality Graphical abstract

scholarly journals Rapid and sensitive detection of SARS-CoV-2 antibodies by biolayer interferometry

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
John V. Dzimianski ◽  
Nicholas Lorig-Roach ◽  
Sara M. O’Rourke ◽  
David L. Alexander ◽  
Jacqueline M. Kimmey ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Optical Measurements ◽  
Enzyme Linked Immunosorbent Assay ◽  
High Complexity ◽  
Antibody Isotype ◽  
Biolayer Interferometry ◽  
Plasma Antibody ◽  
Quantitative Results ◽  
Lateral Flow Test ◽  
Antibody Levels

AbstractSerological testing to evaluate antigen-specific antibodies in plasma is generally performed by rapid lateral flow test strips that lack quantitative results or by high complexity immunoassays that are time- and labor-intensive but provide semi-quantitative results. Here, we describe a novel application of biolayer interferometry for the rapid detection of antigen-specific antibody levels in plasma samples, and demonstrate its utility for quantification of SARS-CoV-2 antibodies. Our biolayer interferometry immunosorbent assay (BLI-ISA) utilizes single-use biosensors in an automated “dip-and-read” format, providing real-time optical measurements of antigen loading, plasma antibody binding, and antibody isotype detection. Complete semi-quantitative results are obtained in less than 20 min. BLI-ISA meets or exceeds the performance of high complexity methods such as Enzyme-Linked Immunosorbent Assay (ELISA) and Chemiluminescent Immunoassay. Importantly, our method can be immediately implemented on existing BLI platforms for urgent COVID-19 studies, such as serosurveillance and the evaluation of vaccine candidates. In a broader sense, BLI-ISA can be developed as a novel diagnostic platform to evaluate antibodies and other biomolecules in clinical specimens.

scholarly journals Potency determination of inactivated H7 influenza vaccines using monoclonal antibody-based ELISA and biolayer interferometry assays

2017 ◽  
Vol 12 (2) ◽  
pp. 250-258 ◽  
Author(s):  
Anupama Vasudevan ◽  
Amy Woerner ◽  
Falko Schmeisser ◽  
Swati Verma ◽  
Ollie Williams ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Influenza Vaccines ◽  
Biolayer Interferometry

scholarly journals Monitoring the Kinetics of the pH-Driven Transition of the Anthrax Toxin Prepore to the Pore by Biolayer Interferometry and Surface Plasmon Resonance

Biochemistry ◽  
2013 ◽  
Vol 52 (37) ◽  
pp. 6335-6347 ◽  
Author(s):  
Subhashchandra Naik ◽  
Susan Brock ◽  
Narahari Akkaladevi ◽  
Jon Tally ◽  
Wesley Mcginn-Straub ◽  
...  
Keyword(s):  
Surface Plasmon Resonance ◽  
Surface Plasmon ◽  
Plasmon Resonance ◽  
Anthrax Toxin ◽  
Biolayer Interferometry ◽  
Kinetics Of

scholarly journals Biolayer interferometry of lipid nanodisc-reconstituted yeast vacuolar H+-ATPase

2017 ◽  
Vol 26 (5) ◽  
pp. 1070-1079 ◽  
Author(s):  
Stuti Sharma ◽  
Stephan Wilkens
Keyword(s):  
Biolayer Interferometry

Characterization of antibody-C1q interactions by Biolayer Interferometry

2018 ◽  
Vol 549 ◽  
pp. 143-148 ◽  
Author(s):  
Wei Zhou ◽  
Shanshan Lin ◽  
Rongying Chen ◽  
Jun Liu ◽  
Yali Li
Keyword(s):  
Biolayer Interferometry

scholarly journals Binding of Brucella protein, Bp26, to select extracellular matrix molecules

2019 ◽  
Vol 20 (1) ◽  
Author(s):  
Yasmin ElTahir ◽  
Amna Al-Araimi ◽  
Remya R. Nair ◽  
Kaija J. Autio ◽  
Hongmin Tu ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Type I Collagen ◽  
Host Cells ◽  
Mouse Serum ◽  
Enzyme Linked Immunosorbent Assay ◽  
Target Cells ◽  
Type I ◽  
Dependent Manner ◽  
Biolayer Interferometry

Abstract Background Brucella is a facultative intracellular pathogen responsible for zoonotic disease brucellosis. Little is known about the molecular basis of Brucella adherence to host cells. In the present study, the possible role of Bp26 protein as an adhesin was explored. The ability of Brucella protein Bp26 to bind to extracellular matrix (ECM) proteins was determined by enzyme-linked immunosorbent assay (ELISA) and biolayer interferometry (BLI). Results ELISA experiments showed that Bp26 bound in a dose-dependent manner to both immobilized type I collagen and vitronectin. Bp26 bound weakly to soluble fibronectin but did not bind to immobilized fibronectin. No binding to laminin was detected. Biolayer interferometry showed high binding affinity of Bp26 to immobilized type I collagen and no binding to fibronectin or laminin. Mapping of Bp26 antigenic epitopes by biotinylated overlapping peptides spanning the entire sequence of Bp26 using anti Bp26 mouse serum led to the identification of five linear epitopes. Collagen and vitronectin bound to peptides from several regions of Bp26, with many of the binding sites for the ligands overlapping. The strongest binding for anti-Bp26 mouse serum, collagen and vitronectin was to the peptides at the C-terminus of Bp26. Fibronectin did not bind to any of the peptides, although it bound to the whole Bp26 protein. Conclusions Our results highlight the possible role of Bp26 protein in the adhesion process of Brucella to host cells through ECM components. This study revealed that Bp26 binds to both immobilized and soluble type I collagen and vitronectin. It also binds to soluble but not immobilized fibronectin. However, Bp26 does not bind to laminin. These are novel findings that offer insight into understanding the interplay between Brucella and host target cells, which may aid in future identification of a new target for diagnosis and/or vaccine development and prevention of brucellosis.

Development of anti-Müllerian hormone immunoassay based on biolayer interferometry technology

2019 ◽  
Vol 411 (21) ◽  
pp. 5499-5507 ◽  
Author(s):  
Yu Wang ◽  
Emmanuel Enoch Dzakah ◽  
Ye Kang ◽  
Yanxue Cai ◽  
Peidian Wu ◽  
...  
Keyword(s):  
Biolayer Interferometry
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