Highly specific label-free protein detection from lysed cells using internally referenced microcantilever sensors

被引:38
作者
Shu, Wenmiao [1 ]
Laurenson, Sophie [2 ]
Knowles, Tuomas P. J. [1 ]
Ferrigno, Paul Ko [2 ]
Seshia, Ashwin A. [1 ,3 ]
机构
[1] Univ Cambridge, Nanosci Ctr, Cambridge CB3 0FF, England
[2] Hutchinson MRC Res Ctr, MRC Canc Cell Unit, Cambridge CB2 2XZ, England
[3] Univ Cambridge, Dept Engn, Cambridge CB2 1PZ, England
基金
英国医学研究理事会; 英国工程与自然科学研究理事会;
关键词
microcantilevers; peptide aptamers; protein fishing;
D O I
10.1016/j.bios.2008.03.036
中图分类号
Q6 [生物物理学];
学科分类号
071011 ;
摘要
We report the investigation of label-free protein detection directly from lysed cells using microcantilever sensors. The integration of an internally referenced microcantilever sensor combined with peptide aptamer technology enables scalable and label-free detection of proteins from a complex biological environment (e.g. cell lysate). The internally referenced microcantilever sensor was found to be effective in minimizing both the effects of thermal drift and non-specific binding interactions with the backside of the cantilever, thereby allowing protein detection in a complex biological background. Highly specific peptide aptamers are used to modify the cantilever surface to specifically detect less than 80 nM CDK2 protein from yeast cell lysate. This binding of CDK2 on the microcantilever generates a tensile surface stress of average magnitude equal to 70 +/- 22 mN/m. Similar experiments conducted with quartz crystal microbalance (QCM) technology are consistent with the response observed using microcantilever sensors. (C) 2008 Elsevier B.V. All rights reserved.
引用
收藏
页码:233 / 237
页数:5
相关论文
共 25 条
[1]   Label-free protein assay based on a nanomechanical cantilever array [J].
Arntz, Y ;
Seelig, JD ;
Lang, HP ;
Zhang, J ;
Hunziker, P ;
Ramseyer, JP ;
Meyer, E ;
Hegner, M ;
Gerber, C .
NANOTECHNOLOGY, 2003, 14 (01) :86-90
[2]   A label-free immunosensor array using single-chain antibody fragments [J].
Backmann, N ;
Zahnd, C ;
Huber, F ;
Bietsch, A ;
Plückthun, A ;
Lang, HP ;
Güntherodt, HJ ;
Hegner, M ;
Gerber, C .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2005, 102 (41) :14587-14592
[3]   Engineering novel binding proteins from nonimmunoglobulin domains [J].
Binz, HK ;
Amstutz, P ;
Plückthun, A .
NATURE BIOTECHNOLOGY, 2005, 23 (10) :1257-1268
[4]   New developments in microarray technology [J].
Blohm, DH ;
Guiseppi-Elie, A .
CURRENT OPINION IN BIOTECHNOLOGY, 2001, 12 (01) :41-47
[5]   Genetic selection of peptide aptamers that recognize and inhibit cyclin-dependent kinase 2 [J].
Colas, P ;
Cohen, B ;
Jessen, T ;
Grishina, I ;
McCoy, J ;
Brent, R .
NATURE, 1996, 380 (6574) :548-550
[6]   Peptide aptamers in label-free protein detection: 1. Characterization of the immobilized scaffold [J].
Davis, Jason J. ;
Tkac, Jan ;
Laurenson, Sophie ;
Ferrigno, Paul Ko .
ANALYTICAL CHEMISTRY, 2007, 79 (03) :1089-1096
[7]   INVITRO SELECTION OF RNA MOLECULES THAT BIND SPECIFIC LIGANDS [J].
ELLINGTON, AD ;
SZOSTAK, JW .
NATURE, 1990, 346 (6287) :818-822
[8]  
ESTOJAK J, 1995, MOL CELL BIOL, V15, P5820
[9]  
Evans David, 2008, J Biol, V7, P3, DOI 10.1186/jbiol62
[10]  
GOLDSBY RA, 2003, IMMUNOLOGY, P148