Improve Research Reproducibility A Bio-protocol resource
Concise Method
tooltip

2.5.3. Competitive FP assays of peptides

XY Xinjian Yin
LC Litong Chen
SY Siwen Yuan
LL Lan Liu
ZG Zhizeng Gao
request Request a Protocol
ask Ask a question
Favorite

Assays were performed manually in black 96 microplates (Cat. No. 3694, Corning, USA) contained 8 nM NusA-5HB and increasing concentrations of peptides (0, 0.31, 0.625, 1.25, 2.5, 5, 10, 20, 40, 80,160, 320 nM) in FP buffer (25 mM pH 7.5 PBS, 0.025% NP-40) in a final volume of 90 μL. After 1 h incubation at 37 °C 200 rpm, 10 μL HR2P-FL (5 nM) was added and additional 1 h incubation was followed at 37 °C 200 rpm. The FP signal was measured. The binding fraction (f) of HR2P-FL with NusA-5HB at different concentrations of peptide was calculated using the Eqs. (1), (2). The obtained f values were plotted using GraphPad Prism 8 to calculate the IC50. All experiments were performed in duplicate.

Copyright and License information:  ©2021 Published by Elsevier Inc.
Since January 2020 Elsevier has created a COVID-19 resource centre with free information in English and Mandarin on the novel coronavirus COVID-19. The COVID-19 resource centre is hosted on Elsevier Connect, the company's public news and information website. Elsevier hereby grants permission to make all its COVID-19-related research that is available on the COVID-19 resource centre - including this research content - immediately available in PubMed Central and other publicly funded repositories, such as the WHO COVID database with rights for unrestricted research re-use and analyses in any form or by any means with acknowledgement of the original source. These permissions are granted for free by Elsevier for as long as the COVID-19 resource centre remains active.

Do you have any questions about this protocol?

Post your question to gather feedback from the community. We will also invite the authors of this article to respond.

0/150

tip Tips for asking effective questions

+ Description

Write a detailed description. Include all information that will help others answer your question including experimental processes, conditions, and relevant images.

post Post a Question
0 Q&A