Methods for using multi-sheath flow and on-chip terminating electrode for microfluidic direct-blotting
Devices and methods are provided for the separation and dispensing of material using a microfluidic separation column connected via an exit channel to one or more sheath flow channels. The flow of separated material through the separation column is at least partially driven by a voltage potential between a first electrode within the separation column and a terminating electrode within at least one of the sheath flow channels. The separation column, exit channel, sheath flow channels, and electrodes are all within a single monolithic chip. The presence of an on-chip terminating electrode allows for separated material to be entrained in the sheath fluids and ejected onto a surface that can be non-conductive. The presence of multiple sheath flows allows for sheath flow fluids to have different compositions from one another, while reducing the occurrence of sheath flow fluids entering the separation column.
1. A method of separating and dispensing an analyte, the method comprising:
providing a separation column, a primary sheath flow channel, a secondary sheath flow channel, and an exit channel, wherein the separation column has an input end and an output end, wherein the output end of the separation column is connected to the exit channel, wherein the primary sheath flow channel intersects the exit channel, and wherein the secondary sheath flow channel intersects the primary sheath flow channel;
pumping a primary sheath fluid through the primary sheath flow channel;
passing a secondary sheath fluid through the secondary sheath flow channel;
applying a voltage between the input end of the separation column and the secondary sheath flow channel, wherein the voltage is sufficient to electrophorese an analyte through the separation column and into the exit channel;
entraining the analyte in the pumped primary sheath fluid to form an effluent; and
ejecting the effluent onto a surface.
2. The method of claim 1 further comprising:
wicking the effluent into a membrane affixed to the surface to immobilize the analyte in the membrane.
3. The method of claim 2 further comprising:
affixing the membrane to the surface and immobilizing the analyte in the membrane using a vacuum manifold connected to the surface.
4. The method of claim 1 wherein the secondary sheath fluid is the primary sheath fluid.
5. The method of claim 1 wherein the secondary sheath fluid is less viscous than the primary sheath fluid, and wherein the secondary sheath fluid has a higher flow rate than that of the primary sheath fluid.
6. The method of claim 1 further comprising:
providing a tertiary sheath flow channel, wherein the tertiary sheath flow channel intersects the exit channel at a location between a discharge outlet of the exit channel and the intersection of the primary sheath flow channel and the exit channel;
flowing an organic solvent through the tertiary sheath flow channel; and
mixing the flowed organic solvent with the effluent.
7. The method of claim 6 wherein the mixing includes forming a spray mixture and wherein the method further comprises:
imparting a voltage between the organic solvent and the surface, wherein the ejecting includes electrospraying of the spray mixture.
8. The method of claim 6 further comprising:
independently adjusting flow rates of the primary sheath fluid, the secondary sheath fluid, and the organic solvent so as to control concentrations of the primary sheath fluid, secondary sheath fluid, and organic solvent in the effluent.
9. The method of claim 1 further comprising:
independently introducing one or more analyte modifiers into one or both of the pumped primary fluid and passed secondary sheath fluid; and
modifying the analyte with the one or more analyte modifiers in the exit channel.
10. The method of claim 9 wherein one or more of the analyte modifiers is a protein stain.
11. The method of claim 1 wherein the secondary sheath fluid is passed through the secondary sheath flow channel using a pressure source.
12. The method of claim 1 wherein the surface or exit channel is moved laterally with respect to the one another using a motor.
13. The method of claim 1 wherein the primary sheath fluid comprises a material selected from the group consisting of dextran, agarose gel, acrylamide, polyacrylamide, silica particles, and combinations thereof.
14. The method of claim 1 wherein the secondary sheath fluid comprises a material selected from the group consisting of water, sodium dodecyl sulfate, a buffer, agarose gel, acrylamide, polyacrylamide, silica particles, an organic solvent, and combinations thereof.