Capillary electrophoresis using microfluidic, electrophoretic, and optic modules
We present here the Lego CE system consisting of available ready-to-use electrophoretic and microfluidic modules, including pressure-based flow controllers. The instrument is coupled with a laser-induced fluorescence detector (LIF) and is demonstrated for separations of labeled oligosaccharides.
We kindly thank the Laboratory of Proteins and Nanotechnology in Analytical Science (PNAS, Institut Galien Paris Saclay, University Paris Saclay) for this collaboration, and for sharing a part of the results obtained with their system.
Materials and Methods
- (Laser-Induced Fluorescence Detector) LIF module
- Electrophoresis module
- Background electrolyte
Figure 2. Electropherograms for CE-LIF separations of oligosaccharide ladders using the Lego CE-LIF instrument; A) Without pressure assistance; B) With pressure assistance at 30 mbar from t = 0s; C) With pressure gradient: 30 mbar at t = 0s, then 20 mbar at t = 5 min.
Separation and detection of APTS labelled oligosaccharides
Glucose-oligosaccharides are often used as the ladder reference for analyzing N-glycans released from glycoproteins, serving for quality control of therapeutic glycoproteins and diagnostic purposes2,3. The Lego CE-LIF was used for separations of APTS-labelled oligosaccharides. Figure 3 shows the electropherograms obtained without pressure assistance (figure 3A), using pressure assistance of 30 mbar t = 0 s (figure 3B), and using pressure assistance of 30 mbar at t=0 s and 20 mbar at t = 5 min (figure 3C). We can observe that excellent peak shapes and separation resolutions are achieved for glucose units GU1 till GU6. To compensate for the peak retardation when using beta-alanine/MES BGE, pressure assistance can be applied during electrophoresis, which is not difficult when using the Flow EZ pressure controller. As we can see in figure 3B, the peaks arrived faster to the detector and more glucose units could be visualized under the pressure assistance at 30 mbar. To finely tune the electrophoresis, it is also possible to use a pressure gradient. By applying a pressure of 30 mbar at 0s and then 20 mbar at 5 min, the fast arrival of the first four peaks could be maintained, whereas separation resolution for the slower ones, which could correspond to the sizes of large N-glycans of glycoproteins, was improved (see figure 3C).
A new Lego CE, that can be fully constructed using commercially-available products, was successfully developed. The need for electronic and mechanical skills, which is often a barrier for constructing in-house CE, is remarkably reduced. The great functioning of this system was demonstrated by separating and detecting fluorescent oligosaccharides. By using pressure assistance provided by the Fluigent instruments, it is possible to accelerate the detection, while keeping a good separation between each peak. Additional experiments have been performed using this system, including testing of different electrolyte, comparing the performance with a commercial CE system. All these results and more information concerning this Lego CE system can be found in the great paper written by Liénard-Mayor et. al1. The Lego design would allow the users to setup their own analytical devices at a cost at least 70 % cheaper than the purchase price of a commercial system while keeping a high degree of standardization (i.e. a ‘standard’ setup) and facilitation of technology transfer that are not offered by in-house-made versions.
- Théo Liénard-Mayor, Jasmine S. Furter, Myriam Taverna, Hung Viet Pham, Peter C. Hauser, T. D. M. Modular instrumentation for capillary electrophoresis with laser induced fluorescence detection using plug-and-play microfluidic, electrophoretic and optic modules. Anal. Chim. Acta 1135, (2020).
- Zhang, P. et al. Challenges of glycosylation analysis and control: An integrated approach to producing optimal and consistent therapeutic drugs. Drug Discov. Today 21, 740–765 (2016).
- Hu, M., Lan, Y., Lu, A., Ma, X. & Zhang, L. Glycan-based biomarkers for diagnosis of cancers and other diseases: Past, present, and future. Progress in Molecular Biology and Translational Science vol. 162 (Elsevier Inc., 2019).