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Issue 34, 2017
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Gradient-free determination of isoelectric points of proteins on chip

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Abstract

The isoelectric point (pI) of a protein is a key characteristic that influences its overall electrostatic behaviour. The majority of conventional methods for the determination of the isoelectric point of a molecule rely on the use of spatial gradients in pH, although significant practical challenges are associated with such techniques, notably the difficulty in generating a stable and well controlled pH gradient. Here, we introduce a gradient-free approach, exploiting a microfluidic platform which allows us to perform rapid pH change on chip and probe the electrophoretic mobility of species in a controlled field. In particular, in this approach, the pH of the electrolyte solution is modulated in time rather than in space, as in the case for conventional determinations of the isoelectric point. To demonstrate the general approachability of this platform, we have measured the isoelectric points of representative set of seven proteins, bovine serum albumin, β-lactoglobulin, ribonuclease A, ovalbumin, human transferrin, ubiquitin and myoglobin in microlitre sample volumes. The ability to conduct measurements in free solution thus provides the basis for the rapid determination of isoelectric points of proteins under a wide variety of solution conditions and in small volumes.

Graphical abstract: Gradient-free determination of isoelectric points of proteins on chip

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Publication details

The article was received on 08 Mar 2017, accepted on 14 Jun 2017 and first published on 17 Aug 2017


Article type: Paper
DOI: 10.1039/C7CP01503H
Citation: Phys. Chem. Chem. Phys., 2017,19, 23060-23067
  • Open access: Creative Commons BY license
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    Gradient-free determination of isoelectric points of proteins on chip

    U. Łapińska, K. L. Saar, E. V. Yates, T. W. Herling, T. Müller, P. K. Challa, C. M. Dobson and T. P. J. Knowles, Phys. Chem. Chem. Phys., 2017, 19, 23060
    DOI: 10.1039/C7CP01503H

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