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Analyst. 2013 Aug 21;138(16):4663-9. doi: 10.1039/c3an00976a. Epub 2013 Jun 19.

Ultra-high density protein spots achieved by on chip digitalized protein synthesis.

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Institute of Industrial Science, The University of Tokyo, 4-6-1, Komaba, Meguro-ku, Tokyo 153-8505, Japan.


Current methodologies for arraying proteins using cell-free protein synthesis on a chip have spatial limitations that prevent reaching ultra-high density necessary for high throughput analysis. To circumvent this, we developed an on-chip method based on microcompartmentalization of protein synthesis. Proteins are synthesized in arrayed micrometer scale chambers from confined DNA template molecules. On-chip protein expression is highly efficient and the method can be used with a minimal amount of template i.e. single DNA molecules to perform digitalized cell-free protein synthesis (d-CFPS). A functionalized surface at the floor of the tightly sealed microchambers enables direct capture of expressed proteins. A density of 10⁴ spots per mm² was achieved, which represents a gain by more than 3 orders of magnitude over conventional methods. This technique of forming such densely arrayed small protein spots is the first step towards the development of a general method that would allow fabrication of ultra-high density protein arrays for high-throughput analysis.

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