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Increasing robustness and sensitivity of protein microarrays through microagitation and automation
NMI Natural and Medical Sciences Institute at the University of Tuebingen.
NMI Natural and Medical Sciences Institute at the University of Tuebingen.
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2006 (English)In: Analytica Chimica Acta, ISSN 0003-2670, E-ISSN 1873-4324, Vol. 564, no 1, 66-73 p.Article in journal (Refereed) Published
Abstract [en]

Assay systems that employ protein microarrays for the analysis of complex samples are powerful tools to generate a high amount of data from a limiting amount of sample. Due to miniaturization, these systems are susceptible to fluctuations during signal generation and the use of uniform conditions for sample incubation and during the assay procedure is required to get reproducible results. Diffusion limits may prevent constant conditions on all parts of the array and can lead to the decease of the sensitivity of the array. Therefore, we set-up an automated assay system integrating a novel microagitation device using surface acoustic wave (SAW) technology. Multiplexed assays for the detection of autoantibodies from human serum and sandwich immunoassay for the detection of matrix metalloproteases (MMPs) were used to evaluate the system. Diffusion-rate limited solid phase reactions were enhanced by microagitation using the SAW technology resulting in up to three-fold higher signals.

Place, publisher, year, edition, pages
2006. Vol. 564, no 1, 66-73 p.
Keyword [en]
automation, surface acoustic waves (SAW), microagitation, protein microarrays, diffusion limitation
National Category
Analytical Chemistry
Identifiers
URN: urn:nbn:se:kth:diva-25837DOI: 10.1016/j.aca.2006.01.041ISI: 000236640100009OAI: oai:DiVA.org:kth-25837DiVA: diva2:360058
Note
QC 20101102Available from: 2010-11-02 Created: 2010-11-02 Last updated: 2017-12-12Bibliographically approved
In thesis
1. Microfluidic Methods for Protein Microarrays
Open this publication in new window or tab >>Microfluidic Methods for Protein Microarrays
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Protein microarray technology has an enormous potential for in vitro diagnostics (IVD)1. Miniaturized and parallelized immunoassays are powerful tools to measure dozens of parameters from minute amounts of sample, whilst only requiring small amounts of reagent. Protein microarrays have become well-established research tools in basic and applied research and the first diagnostic products are already released on the market. However, in order for protein microarrays to become broadly accepted tools in IVD, a number of criteria have to be fulfilled concerning robustness and automation. Robustness and automation are key demands to improve assay performance and reliability of multiplexed assays, and to minimize the time of analysis.

These key demands are addressed in this thesis and novel methods and techniques concerning assay automation, array fabrication as well as performance and detection strategies related to protein microarrays are presented and discussed. In the first paper an automated assay format, based on planar protein microarrays is described and evaluated by the detection of several auto-antibodies from human serum and by quantification of matrix metalloproteases present in plasma. Diffusion-rate limited solid phase reactions were enhanced by microagitation, using the surface acoustic wave technology, resulting in a slightly increased signal-to-noise ratio. In the second paper of the thesis, a novel multiplexed immunoassay system was developed by combining a direct immunoassay with a competitive system. This set-up allows quantification of analytes present in widely varying concentrations within a single multiplex assay. In the third paper, a new concept for sample deposition is introduced, addressing contemporary problems of contact or non-contact microarrayers in protein microarray fabrication.

In the fourth paper, a magnetic bead-based detection method for protein microarrays is described as a cost-effective alternative approach to the commonly used fluorescence-based confocal scanning systems. The magnetic bead-based detection could easily be performed by using an ordinary flatbed scanner. In addition, applying magnetic force to the magnetic bead-based detection approach enables to run the detection step more rapidly. Finally, in paper five, a microfluidic bead-based immunoassay for multiplexed detection of receptor tyrosine kinases in breast cancer tissue is presented. Since the assay is performed inside a capillary, the amounts of sample and reagent material could be reduced by a factor of 30 or more when compared with the current standard protein microarray assay.

Place, publisher, year, edition, pages
Stockholm: KTH, 2010. ix, 51 p.
Series
Trita-CHE-Report, ISSN 1654-1081 ; 2010:45
National Category
Analytical Chemistry
Identifiers
urn:nbn:se:kth:diva-26083 (URN)978-91-7415-761-1 (ISBN)
Public defence
2010-11-19, F3, Lindstedtsvägen 26, KTH, Stockholm, 10:00 (English)
Opponent
Supervisors
Note
QC 20101112Available from: 2010-11-12 Created: 2010-11-12 Last updated: 2010-11-15Bibliographically approved

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