57862 20180515100241.0 doi 10.1371/journal.pone.0102572 sideral 91898 ART-2014-91898 eng Chakraborty, Sandeep FRET based quantification and screening technology platform for the interactions of Leukocyte Function-associated Antigen-1 (LFA-1) with InterCellular Adhesion Molecule-1 (ICAM-1) 2014 Access copy available to the general public Unrestricted The interaction between leukocyte function-associated antigen-1(LFA-1) and intercellular adhesion molecule-1 (ICAM-1) plays a pivotal role in cellular adhesion including the extravasation and inflammatory response of leukocytes, and also in the formation of immunological synapse. However, irregular expressions of LFA-1 or ICAM-1 or both may lead to autoimmune diseases, metastasis cancer, etc. Thus, the LFA-1/ICAM-1 interaction may serve as a potential therapeutic target for the treatment of these diseases. Here, we developed one simple ‘in solution’ steady state fluorescence resonance energy transfer (FRET) technique to obtain the dissociation constant (Kd) of the interaction between LFA-1 and ICAM-1. Moreover, we developed the assay into a screening platform to identify peptides and small molecules that inhibit the LFA-1/ICAM-1 interaction. For the FRET pair, we used Alexa Fluor 488-LFA-1 conjugate as donor and Alexa Fluor 555-human recombinant ICAM-1 (D1-D2-Fc) as acceptor. From our quantitative FRET analysis, the Kd between LFA-1 and D1-D2-Fc was determined to be 17.93±1.34 nM. Both the Kd determination and screening assay were performed in a 96-well plate platform, providing the opportunity to develop it into a high-throughput assay. This is the first reported work which applies FRET based technique to determine Kd as well as classifying inhibitors of the LFA-1/ICAM-1 interaction. info:eu-repo/grantAgreement/ES/MINECO-FEDER/SAF2011-25390 info:eu-repo/semantics/openAccess by http://creativecommons.org/licenses/by/3.0/es/ 3.234 2014 MULTIDISCIPLINARY SCIENCES 9 / 57 = 0.158 2014 Q1 T1 info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion Núñez, David Hu, Shih-Yang Domingo, María Pilar (orcid)0000-0003-0154-0730 Pardo, Julian Universidad de Zaragoza Karmenyan, Artashes Gálvez, Eva M Chiou, Arthur 1008 566 Universidad de Zaragoza Departamento de Microbiología, Medicina Preventiva y Salud Pública Inmunología 9, 7 (2014), e102572 [13 pp] PLoS One PLoS One 1932-6203 628023 http://zaguan.unizar.es/record/57862/files/texto_completo.pdf Versión publicada 122121 http://zaguan.unizar.es/record/57862/files/texto_completo.jpg?subformat=icon icon Versión publicada oai:zaguan.unizar.es:57862 articulos driver 2018-05-15-09:59:17 ARTICLE