Subnanoliter enzymatic assays on microarrays

Many areas of research today are based on enzymatic assays most of which are still performed as enzyme‐linked immunosorbent assays in microtiter plates. The demand for highly parallel screening of thousands of samples eventually led to a miniaturization and automation of these assays. However, the f...

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Bibliographic Details
Published in:Proteomics (Weinheim) 2005-02, Vol.5 (2), p.420-425
Main Authors: Angenendt, Philipp, Lehrach, Hans, Kreutzberger, Jürgen, Glökler, Jörn
Format: Article
Language:English
Subjects:
Alkaline Phosphatase - analysis
Alkaline Phosphatase - antagonists & inhibitors
Analytical, structural and metabolic biochemistry
beta-Galactosidase - analysis
beta-Galactosidase - antagonists & inhibitors
Biological and medical sciences
Cathepsin D - analysis
Drug screening
Enzymatic assay
Enzymes - analysis
Fundamental and applied biological sciences. Psychology
Horseradish Peroxidase - analysis
Horseradish Peroxidase - antagonists & inhibitors
Metagenomics
Miscellaneous
Multiple spotting technique
Nanotechnology
Polystyrenes - chemistry
Protein Array Analysis
Protein microarray
Proteins
Proteomics
Sensitivity and Specificity
Surface Properties
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Summary:Many areas of research today are based on enzymatic assays most of which are still performed as enzyme‐linked immunosorbent assays in microtiter plates. The demand for highly parallel screening of thousands of samples eventually led to a miniaturization and automation of these assays. However, the final transfer of enzymatic assays from a microtiter‐based technology to microarrays has proven to be difficult for various reasons, such as the inability to maintain unbound reaction products on the spot of reaction or the missing capability of multiplexing. Here, we have conducted multiplex enzymatic assays in subnanoliter volumes on a single microarray using the multiple spotting technology. We were able to measure enzymatic activity with a sensitivity down to 35 enzyme molecules, applying only conventional flat microarray surfaces and standard microarray hardware. We have performed assays of inhibition and applied this format for the detection of prognostic markers, such as cathepsin D. The new approach allows the rapid and multiplex screening of thousands of samples on a single microarray with applications in drug screening, metagenomics, and high‐throughput enzyme assays.
ISSN:1615-9853
1615-9861
DOI:10.1002/pmic.200400955