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Anal Biochem. 2009 Mar 15;386(2):194-216. doi: 10.1016/j.ab.2008.11.021. Epub 2008 Nov 27.

A global benchmark study using affinity-based biosensors.

Rich RL1, Papalia GA, Flynn PJ, Furneisen J, Quinn J, Klein JS, Katsamba PS, Waddell MB, Scott M, Thompson J, Berlier J, Corry S, Baltzinger M, Zeder-Lutz G, Schoenemann A, Clabbers A, Wieckowski S, Murphy MM, Page P, Ryan TE, Duffner J, Ganguly T, Corbin J, Gautam S, Anderluh G, Bavdek A, Reichmann D, Yadav SP, Hommema E, Pol E, Drake A, Klakamp S, Chapman T, Kernaghan D, Miller K, Schuman J, Lindquist K, Herlihy K, Murphy MB, Bohnsack R, Andrien B, Brandani P, Terwey D, Millican R, Darling RJ, Wang L, Carter Q, Dotzlaf J, Lopez-Sagaseta J, Campbell I, Torreri P, Hoos S, England P, Liu Y, Abdiche Y, Malashock D, Pinkerton A, Wong M, Lafer E, Hinck C, Thompson K, Primo CD, Joyce A, Brooks J, Torta F, Bagge Hagel AB, Krarup J, Pass J, Ferreira M, Shikov S, Mikolajczyk M, Abe Y, Barbato G, Giannetti AM, Krishnamoorthy G, Beusink B, Satpaev D, Tsang T, Fang E, Partridge J, Brohawn S, Horn J, Pritsch O, Obal G, Nilapwar S, Busby B, Gutierrez-Sanchez G, Gupta RD, Canepa S, Witte K, Nikolovska-Coleska Z, Cho YH, D'Agata R, Schlick K, Calvert R, Munoz EM, Hernaiz MJ, Bravman T, Dines M, Yang MH, Puskas A, Boni E, Li J, Wear M, Grinberg A, Baardsnes J, Dolezal O, Gainey M, Anderson H, Peng J, Lewis M, Spies P, Trinh Q, Bibikov S, Raymond J, Yousef M, Chandrasekaran V, Feng Y, Emerick A, Mundodo S, Guimaraes R, McGirr K, Li YJ, Hughes H, Mantz H, Skrabana R, Witmer M, Ballard J, Martin L, Skladal P, Korza G, Laird-Offringa I, Lee CS, Khadir A, Podlaski F, Neuner P, Rothacker J, Rafique A, Dankbar N, Kainz P, Gedig E, Vuyisich M, Boozer C, Ly N, Toews M, Uren A, Kalyuzhniy O, Lewis K, Chomey E, Pak BJ, Myszka DG.

Author information

1
Center for Biomolecular Interaction Analysis, School of Medicine, University of Utah, Salt Lake City, UT 84132, USA.

Abstract

To explore the variability in biosensor studies, 150 participants from 20 countries were given the same protein samples and asked to determine kinetic rate constants for the interaction. We chose a protein system that was amenable to analysis using different biosensor platforms as well as by users of different expertise levels. The two proteins (a 50-kDa Fab and a 60-kDa glutathione S-transferase [GST] antigen) form a relatively high-affinity complex, so participants needed to optimize several experimental parameters, including ligand immobilization and regeneration conditions as well as analyte concentrations and injection/dissociation times. Although most participants collected binding responses that could be fit to yield kinetic parameters, the quality of a few data sets could have been improved by optimizing the assay design. Once these outliers were removed, the average reported affinity across the remaining panel of participants was 620 pM with a standard deviation of 980 pM. These results demonstrate that when this biosensor assay was designed and executed appropriately, the reported rate constants were consistent, and independent of which protein was immobilized and which biosensor was used.

PMID:
19133223
PMCID:
PMC3793259
DOI:
10.1016/j.ab.2008.11.021
[Indexed for MEDLINE]
Free PMC Article

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