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Novel concept microarray enabling PCR and multistep reactions through pipette-free aperture-to-aperture parallel transfer.

Kinoshita Y, Tayama T, Kitamura K, Salimullah M, Uchida H, Suzuki M, Husimi Y, Nishigaki K - BMC Biotechnol. (2010)

Bottom Line: On the other hand, the popular microplate technology, which has a great merit of being able to perform parallel multistep reactions, has come to its limit in increasing the number of wells (currently, up to 9600) and reducing the volume to deal with due to the difficulty in operations.These were demonstrated by applying the MMV technology to searching lysozyme-crystallizing conditions and selecting peptides aimed for Aβ-binding or cathepsin E-inhibition.With the introduction of a novel concept microarray (MMV) technology, parallel and multistep reactions in sub-μL scale have become possible.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Functional Materials Science, Graduate School of Science and Engineering, Saitama University, 255 Shimo-okubo, Saitama 338-8570, Japan.

ABSTRACT

Background: The microarray has contributed to developing the omic analysis. However, as it depends basically on the surface reaction, it is hard to perform bulk reactions and sequential multistep reactions. On the other hand, the popular microplate technology, which has a great merit of being able to perform parallel multistep reactions, has come to its limit in increasing the number of wells (currently, up to 9600) and reducing the volume to deal with due to the difficulty in operations.

Results: Here, we report a novel microarray technology which enables us to explore advanced applications, termed microarray-with-manageable volumes (MMV). The technical essence is in the pipette-free direct parallel transfer from well to well performed by centrifugation, evading the evaporation and adsorption-losses during handling. By developing the MMV plate, accompanying devices and techniques, generation of multiple conditions (256 kinds) and performance of parallel multistep reactions, including PCR and in vitro translation reactions, have been made possible. These were demonstrated by applying the MMV technology to searching lysozyme-crystallizing conditions and selecting peptides aimed for Aβ-binding or cathepsin E-inhibition.

Conclusions: With the introduction of a novel concept microarray (MMV) technology, parallel and multistep reactions in sub-μL scale have become possible.

Show MeSH
MMV cultivation of E. coli. GFP-expressing cells of E. coli were detected by fluorescence with a fluoroimager (FITC mode): Dark wells indicate fluorescent colonies of E. coli.
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Figure 8: MMV cultivation of E. coli. GFP-expressing cells of E. coli were detected by fluorescence with a fluoroimager (FITC mode): Dark wells indicate fluorescent colonies of E. coli.

Mentions: In addition to the plastic MMV made of PDMS (polydimethylsiloxane) or acrylate resin as a dry type, we originally introduced a wet type MMV made of polyacrylamide gel. To validate wet MMVs used for cultivation, bacteria E. coli harboring green fluorescent protein (GFP) were subjected to a single cell culture (i.e., a single cell per well (expectation value)). Overnight culture of the cells in the MMV resulted in a Poisson distribution as expected (Fig. 8). This means that we could obtain neatly arrayed colonies without depending on robotics such as Colony Picker [33], which can be easily manipulated in the following processes.


Novel concept microarray enabling PCR and multistep reactions through pipette-free aperture-to-aperture parallel transfer.

Kinoshita Y, Tayama T, Kitamura K, Salimullah M, Uchida H, Suzuki M, Husimi Y, Nishigaki K - BMC Biotechnol. (2010)

MMV cultivation of E. coli. GFP-expressing cells of E. coli were detected by fluorescence with a fluoroimager (FITC mode): Dark wells indicate fluorescent colonies of E. coli.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2959086&req=5

Figure 8: MMV cultivation of E. coli. GFP-expressing cells of E. coli were detected by fluorescence with a fluoroimager (FITC mode): Dark wells indicate fluorescent colonies of E. coli.
Mentions: In addition to the plastic MMV made of PDMS (polydimethylsiloxane) or acrylate resin as a dry type, we originally introduced a wet type MMV made of polyacrylamide gel. To validate wet MMVs used for cultivation, bacteria E. coli harboring green fluorescent protein (GFP) were subjected to a single cell culture (i.e., a single cell per well (expectation value)). Overnight culture of the cells in the MMV resulted in a Poisson distribution as expected (Fig. 8). This means that we could obtain neatly arrayed colonies without depending on robotics such as Colony Picker [33], which can be easily manipulated in the following processes.

Bottom Line: On the other hand, the popular microplate technology, which has a great merit of being able to perform parallel multistep reactions, has come to its limit in increasing the number of wells (currently, up to 9600) and reducing the volume to deal with due to the difficulty in operations.These were demonstrated by applying the MMV technology to searching lysozyme-crystallizing conditions and selecting peptides aimed for Aβ-binding or cathepsin E-inhibition.With the introduction of a novel concept microarray (MMV) technology, parallel and multistep reactions in sub-μL scale have become possible.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Functional Materials Science, Graduate School of Science and Engineering, Saitama University, 255 Shimo-okubo, Saitama 338-8570, Japan.

ABSTRACT

Background: The microarray has contributed to developing the omic analysis. However, as it depends basically on the surface reaction, it is hard to perform bulk reactions and sequential multistep reactions. On the other hand, the popular microplate technology, which has a great merit of being able to perform parallel multistep reactions, has come to its limit in increasing the number of wells (currently, up to 9600) and reducing the volume to deal with due to the difficulty in operations.

Results: Here, we report a novel microarray technology which enables us to explore advanced applications, termed microarray-with-manageable volumes (MMV). The technical essence is in the pipette-free direct parallel transfer from well to well performed by centrifugation, evading the evaporation and adsorption-losses during handling. By developing the MMV plate, accompanying devices and techniques, generation of multiple conditions (256 kinds) and performance of parallel multistep reactions, including PCR and in vitro translation reactions, have been made possible. These were demonstrated by applying the MMV technology to searching lysozyme-crystallizing conditions and selecting peptides aimed for Aβ-binding or cathepsin E-inhibition.

Conclusions: With the introduction of a novel concept microarray (MMV) technology, parallel and multistep reactions in sub-μL scale have become possible.

Show MeSH