First Separation using the microfluidic DFGF

The first experiment attempted with the new device was a separation mixture of 10%v/v BPB and AM prepared in 50mM tris HCl buffer solution. The flow rate was set at 1µLmin-1 and 10µL of the test mixture was injected.

98 Once the sample entered the separation channel, and the two components began to resolve, the analytes came into poorly focused bands near the beginning of the channel, figure 3.3.4. After ~20mins the sample began to migrate further along the channel and eluted from the device with no changes made to the EFG. The conditions applied are shown in table 3.3.1.

Time (mins) EFG (Vcm-2_{) and Voltage Profile Flow(µLmin}-1₎

1

G=100

1.0

99 Figure 3.3.4. Initial Separation of BPB and AM. A) Raw image of the separation B) Enlarged view of the separation with focused bands of AM and BPB. C) Graphical representation of the separation. The peaks come into focus before fading as a result of loss of EFG.

A B

100 3.3.4 Electrode decomposition, New Gold electrodes

Having first ensured there were no issues with the system configuration and the power supply was indeed functioning, the experiment was repeated. The repeat experiment was conducted using the same conditions (table 3.3.1). From the observation of the injected sample passing through the separation channel with no retention or focusing, it was clear there was a serious problem with the device itself. On further examination, it was immediately apparent that the electrode chip was the source of the device failure. The platinum electrodes which had been deposited on the surface had significantly degraded. The degradation occurred as the high current density oxidised the platinum surface removing it from the substrate. The extent of the decomposition is shown in figure 3.3.5.

Figure 3.3.5. Severe degradation of the platinum electrodes on the microfluidic DFGF electrode chip. The area highlighted in red displays the degraded electrodes.

To protect the electrodes, and lower the high quantity of electrolysis gasses produced, an electrode plate was coated with a layer of Nafion. The coating was carried out by sequentially dropping 5%v/v Nafion (Sigma Aldrich) suspension onto the surface and evaporating the solvent (water) by heating to 60°C overnight.

101 Nafion is a fluropoly-copolymer developed in Germany by DuPont in the 1960’s

shown in figure 3.3.621_{. This polymer was considered due to many electrochemical}

applications in electro-catalysis22 _{and as membranes in fuel cells}23 _{as it only} facilitates the permeation of small charged ions. With these properties this polymer was also considered as a viable alternative to the cellulose membrane.

Figure 3.3.6. Molecular structure of Nafion21_.

Using the conditions in table 3.3.1 a two component test mixture of BPB and AM was injected. The microfluidic DFGF was set up with the same configuration used previously, the only difference being the Nafion Coating over the electrodes. As observed in the initial experiment, the analytes began to focus into bands ~10mm along the separation channel. However, the focused bands faded and eluted from the device. It was later determined that the Nafion coating had deteriorated along with another set of Platinum electrodes the same as in figure 3.3.5. With these results came the realisation that an alternative electrode array was required.

102 One of the failed electrode chips was taken and five grooves ~350µm deep were tooled into the surface equally spaced 5mm apart with the 3rd _{electrode centred} between the two drilled holes in the chip. Inlays of 500µm diameter gold wire were secured in these grooves using cyanoacrylate. The exposed surface of gold wire was milled down level to the surface of the electrode chip and the whole surface was then polished. The resulting new gold electrode array is shown in figure 3.3.7.

Figure 3.3.7. Gold electrode array on an existing electrode chip.

Integrating this new electrode array into the microfluidic DFGF device involved soldering the electrodes to another IDE cable with and IDC connector. As the soldered joint to these electrodes was brittle the connector was incorporated into the base. As this was a fixed solution, the ‘Gasket holder’ and electrode channel gasket were set in place and the recess filled with epoxy resin to seal the electrode channel. The completed electrode array and ‘Chip Holder Base’ are shown in figure 3.3.8.

103 Figure 3.3.8. Different views of the completed gold electrode array and chip holder base with fixed connector.

A new test mixture had been formulated with a third component for these experiments to further demonstrate the degree of the dynamic control over the EFG by identifying changes in the field with an additional focal point. This third component, colloquially known as “Acid Yellow”, was added as it has a different hydrodynamic radius with less net charge than BPB and AM. The molecular structure of Acid Yellow (AY) is shown in figure 3.3.9.

10mm 10mm

104 Figure 3.3.9. The molecular structure of Disodium 2,5-dichloro-4-[3-methyl-5-oxo-4-(4- sulfonatophenyl)diazenyl-4H-pyrazol-1-yl]benzenesulfonate (Acid Yellow).

With the new gold electrodes incorporated into the ‘chip holder base’, preparing the device was simplified. The membrane was placed on top of the secured gasket and the separation channel chip inserted into the locating slots before being clamped down over the membrane with the ‘chip holder top’. To complete the setup the two thumb screws were tightened securely.