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On-chip Isotachophoresis for Separation of Ions and Purification of Nucleic Acids

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Two-step perpendicular free-solution isoelectric focusing in a microchamber array chip.

Ryo Ishibashi¹, Takehiko Kitamori, Kiyohito Shimura

¹Department of Applied Chemistry, School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo, Tokyo 113-8656, Japan.

|August 11, 2010

Summary

A novel microfluidic chip enables rapid, small-scale protein fractionation using two-step perpendicular isoelectric focusing (IEF). This device achieves efficient separation of peptides with minimal isoelectric point differences in just 10 minutes.

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Area of Science:

Biochemistry
Analytical Chemistry
Microfluidics

Background:

Isoelectric focusing (IEF) is a powerful protein separation technique.
Traditional gel-based IEF can be time-consuming and requires larger sample volumes.
Developing miniaturized, efficient IEF systems is crucial for high-throughput analysis.

Purpose of the Study:

To design and fabricate a free-solution microfluidic chip for small-scale protein fractionation.
To implement a two-step perpendicular isoelectric focusing (IEF) method on the microchip.
To evaluate the chip's performance in terms of separation speed, voltage, and resolution.

Main Methods:

Fabrication of a microfluidic chip using two glass plates with etched indentations.
Formation of channels or chambers by adjusting the relative position of the glass plates.
Application of two-step perpendicular isoelectric focusing (IEF) for protein separation.
Analysis of fluorescence-labeled peptides with a small isoelectric point difference (DeltapI = 0.3).

Main Results:

The microfluidic chip successfully fractionated proteins based on isoelectric point (pI).
Two-step perpendicular IEF achieved separation into 100 fractions.
Efficient separation of peptides with DeltapI = 0.3 was demonstrated.
Total separation time was reduced to 10 minutes at a maximum voltage of 500 V.

Conclusions:

The developed microfluidic chip offers a rapid and efficient platform for small-scale protein fractionation.
The two-step perpendicular IEF approach on-chip enhances separation resolution and speed.
This technology holds promise for applications requiring fast, high-resolution protein analysis.