Development of a Cell-Based Assay for the Detection of Neutralizing Antibodies to PF-06730512 Using Homogenous Time-Resolved Fluorescence

2020 ◽  
Vol 22 (2) ◽  
Author(s):  
Michael Luong ◽  
Ying Wang ◽  
Stephen P. Berasi ◽  
Janet E. Buhlmann ◽  
Hongying Yang ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Time Resolved Fluorescence ◽  
Time Resolved ◽  
A Cell

A cell-based time-resolved fluorescence assay for selection of antibody reagents for G protein-coupled receptor immunohistochemistry

2004 ◽  
Vol 291 (1-2) ◽  
pp. 123-135 ◽  
Author(s):  
Jui-Lan Su ◽  
Jim Fornwald ◽  
Philip Rivers ◽  
Susan Goldsworthy ◽  
Noeleen A. Looney ◽  
...  
Keyword(s):  
G Protein ◽  
Fluorescence Assay ◽  
Time Resolved Fluorescence ◽  
G Protein Coupled Receptor ◽  
Time Resolved ◽  
A Cell ◽  
G Protein Coupled ◽  
Selection Of

scholarly journals Comparison of Assay Technologies for a Nuclear Receptor Assay Screen Reveals Differences in the Sets of Identified Functional Antagonists

2003 ◽  
Vol 8 (4) ◽  
pp. 381-392 ◽  
Author(s):  
Xiang Wu ◽  
J. Fraser Glickman ◽  
Benjamin R. Bowen ◽  
Matthew A. Sills
Keyword(s):  
Nuclear Receptor ◽  
Dose Response ◽  
High Throughput Screening ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Functional Assay ◽  
Time Resolved Fluorescence ◽  
Time Resolved ◽  
Receptor Assay ◽  
A Cell

Many assay technologies currently exist to develop high-throughput screening assays, and the number of choices continues to increase. Results from a previous study comparing assay technologies in our laboratory do not support the common assumption that the same hits would be found regardless of which assay technology is used. To extend this investigation, a nuclear receptor antagonist assay was developed using 3 assay formats: AlphaScreen, time-resolved fluorescence (TRF), and time-resolved fluorescence resonance energy transfer (TR-FRET). Compounds (∼42,000) from the Novartis library were evaluated in all 3 assay formats. A total of 128 compounds were evaluated in dose-response experiments, and 109 compounds were confirmed active from all 3 formats. The AlphaScreen, TRF, and TR-FRET assay technologies identified 104, 23, and 57 active compounds, respectively, with only 18 compounds active in all 3 assay formats. A total of 128 compounds were evaluated in a cell-based functional assay, and 35 compounds demonstrated activity in this cellular assay. Furthermore, 34, 11, and 16 hits that were originally identified in the dose-response experiment by AlphaScreen, TRF, and TR-FRET assay technologies, respectively, were functionally active. The results of the study indicated that AlphaScreen identified the greatest number of functional antagonists.

scholarly journals The influence of sugar–protein complexes on the thermostability of C-reactive protein (CRP)

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Andreea Lorena Mateescu ◽  
Nicolae-Bogdan Mincu ◽  
Silvana Vasilca ◽  
Roxana Apetrei ◽  
Diana Stan ◽  
...  
Keyword(s):  
Diagnostic Tests ◽  
Protein Complexes ◽  
C Reactive Protein ◽  
Time Resolved Fluorescence ◽  
Time Resolved ◽  
Reactive Protein ◽  
In Vitro Diagnostic ◽  
The Stability ◽  
Positive Controls

AbstractThe purpose of the present study was to evaluate de influence of protein–sugar complexation on the stability and functionality of C-reactive protein, after exposure to constant high temperatures, in order to develop highly stable positive controls for in-vitro diagnostic tests. C-reactive protein is a plasmatic protein used as a biomarker for the diagnosis of a series of health problems such as ulcerative colitis, cardiovascular diseases, metabolic syndrome, due to its essential role in the evolution of chronic inflammation. The sugar–protein interaction was investigated using steady state and time resolved fluorescence. The results revealed that there are more than two classes of tryptophan, with different degree of accessibility for the quencher molecule. Our study also revealed that sugar–protein complexes have superior thermostability, especially after gamma irradiation at 2 kGy, the protein being stable and functional even after 22 days exposure to 40 °C.

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