A global benchmark study using affinity-based biosensors.
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.
Rich
Rebecca L
RL
Center for Biomolecular Interaction Analysis, School of Medicine, University of Utah, Salt Lake City, UT 84132, USA.
Papalia
Giuseppe A
GA
Flynn
Peter J
PJ
Furneisen
Jamie
J
Quinn
John
J
Klein
Joshua S
JS
Katsamba
Phini S
PS
Waddell
M Brent
MB
Scott
Michael
M
Thompson
Joshua
J
Berlier
Judie
J
Corry
Schuyler
S
Baltzinger
Mireille
M
Zeder-Lutz
Gabrielle
G
Schoenemann
Andreas
A
Clabbers
Anca
A
Wieckowski
Sebastien
S
Murphy
Mary M
MM
Page
Phillip
P
Ryan
Thomas E
TE
Duffner
Jay
J
Ganguly
Tanmoy
T
Corbin
John
J
Gautam
Satyen
S
Anderluh
Gregor
G
Bavdek
Andrej
A
Reichmann
Dana
D
Yadav
Satya P
SP
Hommema
Eric
E
Pol
Ewa
E
Drake
Andrew
A
Klakamp
Scott
S
Chapman
Trevor
T
Kernaghan
Dawn
D
Miller
Ken
K
Schuman
Jason
J
Lindquist
Kevin
K
Herlihy
Kara
K
Murphy
Michael B
MB
Bohnsack
Richard
R
Andrien
Bruce
B
Brandani
Pietro
P
Terwey
Danny
D
Millican
Rohn
R
Darling
Ryan J
RJ
Wang
Liann
L
Carter
Quincy
Q
Dotzlaf
Joe
J
Lopez-Sagaseta
Jacinto
J
Campbell
Islay
I
Torreri
Paola
P
Hoos
Sylviane
S
England
Patrick
P
Liu
Yang
Y
Abdiche
Yasmina
Y
Malashock
Daniel
D
Pinkerton
Alanna
A
Wong
Melanie
M
Lafer
Eileen
E
Hinck
Cynthia
C
Thompson
Kevin
K
Primo
Carmelo Di
CD
Joyce
Alison
A
Brooks
Jonathan
J
Torta
Federico
F
Bagge Hagel
Anne Birgitte
AB
Krarup
Janus
J
Pass
Jesper
J
Ferreira
Monica
M
Shikov
Sergei
S
Mikolajczyk
Malgorzata
M
Abe
Yuki
Y
Barbato
Gaetano
G
Giannetti
Anthony M
AM
Krishnamoorthy
Ganeshram
G
Beusink
Bianca
B
Satpaev
Daulet
D
Tsang
Tiffany
T
Fang
Eric
E
Partridge
James
J
Brohawn
Stephen
S
Horn
James
J
Pritsch
Otto
O
Obal
Gonzalo
G
Nilapwar
Sanjay
S
Busby
Ben
B
Gutierrez-Sanchez
Gerardo
G
Gupta
Ruchira Das
RD
Canepa
Sylvie
S
Witte
Krista
K
Nikolovska-Coleska
Zaneta
Z
Cho
Yun Hee
YH
D'Agata
Roberta
R
Schlick
Kristian
K
Calvert
Rosy
R
Munoz
Eva M
EM
Hernaiz
Maria Jose
MJ
Bravman
Tsafir
T
Dines
Monica
M
Yang
Min-Hsiang
MH
Puskas
Agnes
A
Boni
Erica
E
Li
Jiejin
J
Wear
Martin
M
Grinberg
Asya
A
Baardsnes
Jason
J
Dolezal
Olan
O
Gainey
Melicia
M
Anderson
Henrik
H
Peng
Jinlin
J
Lewis
Mark
M
Spies
Peter
P
Trinh
Quyhn
Q
Bibikov
Sergei
S
Raymond
Jill
J
Yousef
Mohammed
M
Chandrasekaran
Vidya
V
Feng
Yuguo
Y
Emerick
Anne
A
Mundodo
Suparna
S
Guimaraes
Rejane
R
McGirr
Katy
K
Li
Yue-Ji
YJ
Hughes
Heather
H
Mantz
Hubert
H
Skrabana
Rostislav
R
Witmer
Mark
M
Ballard
Joshua
J
Martin
Loic
L
Skladal
Petr
P
Korza
George
G
Laird-Offringa
Ite
I
Lee
Charlene S
CS
Khadir
Abdelkrim
A
Podlaski
Frank
F
Neuner
Phillippe
P
Rothacker
Julie
J
Rafique
Ashique
A
Dankbar
Nico
N
Kainz
Peter
P
Gedig
Erk
E
Vuyisich
Momchilo
M
Boozer
Christina
C
Ly
Nguyen
N
Toews
Mark
M
Uren
Aykut
A
Kalyuzhniy
Oleksandr
O
Lewis
Kenneth
K
Chomey
Eugene
E
Pak
Brian J
BJ
Myszka
David G
DG
D017151:Antibodies, Catalytic / Q000032:analysis; D019985:Benchmarking; D001665:Binding Sites; D015374:Biosensing Techniques / Q000379:methods* / Q000706:statistics & numerical data; D005982:Glutathione Transferase / Q000032:analysis; D007700:Kinetics; D008024:Ligands; D011506:Proteins / Q000032:analysis*
D017151:Antibodies, Catalytic; D008024:Ligands; D011506:Proteins; D005982:Glutathione Transferase