Preconcentration and separation of analytes in microchannels
Disclosed herein are methods and devices for preconcentrating and separating analytes such as proteins and polynucleotides in microchannels. As disclosed, at least one size-exclusion polymeric element is adjacent to processing area or an assay area in a microchannel which may be porous polymeric element. The size-exclusion polymeric element may be used to manipulate, e.g. concentrate, analytes in a sample prior to assaying in the assay area.
1. A microfluidic device comprising
a microfluidic channel having
at least one in situ polymerized size-exclusion polymeric element having a first side and a second side, and
at least one porous polymeric element adjacent to the second side,
wherein the size-exclusion polymeric element has a length that is thinner than the length of the porous polymeric element,
a loading channel in fluidic communication with a portion of the microfluidic channel containing the porous polymeric element,
a sample reservoir in fluidic communication with a first waste reservoir, said sample reservoir is configured for containing a sample, wherein the sample reservoir and the first waste reservoir are configured to apply a first electric field from the sample reservoir through the loading channel and from the second side to the first side to the first waste reservoir, and
a first buffer reservoir and a second waste reservoir configured to apply a second electric field from the first buffer reservoir and from the second side to the second waste reservoir through the length of the porous polymeric element, wherein the second electric field bypasses the size-exclusion polymeric element.
2. The microfluidic device of claim 1 , and further comprising a second size-exclusion polymeric element.
3. The microfluidic device of claim 2 , wherein the size-exclusion polymeric element has pores that are larger than the pores of the second size-exclusion polymeric element.
4. The microfluidic device of claim 2 , wherein the size-exclusion polymeric element has a cross-sectional area that is greater than the than the cross-sectional area of the second size-exclusion polymeric element.
5. The microfluidic device of claim 2 , wherein the size-exclusion polymeric elements are arranged in series.
6. The microfluidic device of claim 1 , wherein the size of the pores of the size-exclusion polymeric element are smaller than the size of an analyte of interest in the sample so as to prevent the analyte from passing through the size-exclusion polymeric element under application of the first electric field.
7. The microfluidic device of claim 1 , wherein the length of the size-exclusion polymeric element is about 10 μm to about 50 μm.
8. The microfluidic device of claim 1 , wherein the microfluidic device is configured to apply a third electric field from the first waste reservoir or a second buffer reservoir to the second waste reservoir and from the first side to the second side.
9. A method for processing an analyte in a sample in a microfluidic device comprising a microfluidic channel having at least one in situ polymerized size-exclusion polymeric element having a first side and a second side, and at least one porous polymeric element adjacent to the second side, wherein the size-exclusion polymeric element has a length that is thinner than the length of the porous polymeric element, a loading channel in fluidic communication with a portion of the microfluidic channel containing the porous polymeric element, a sample reservoir in fluidic communication with a first waste reservoir, said sample reservoir is configured for containing a sample, wherein the sample reservoir and the first waste reservoir are configured to apply a first electric field from the sample reservoir through the loading channel and from the second side to the first side to the first waste reservoir, and a first buffer reservoir and a second waste reservoir configured to apply a second electric field from the first buffer reservoir and from the second side to the second waste reservoir through the length of the porous polymeric element, wherein the second electric field bypasses the size-exclusion polymeric element, wherein the method comprises
concentrating the analyte on the second side of the size-exclusion polymeric element, and
moving the analyte through the porous polymeric element.
10. The method of claim 9 , wherein the analyte is concentrated by applying the first electric field.
11. The method of claim 9 , wherein the analyte is moved through the porous polymeric element by applying the second electric field.
12. The method of claim 9 , which further comprises
eluting the analyte off the second side by applying the third electric field.
13. The method of claim 9 , which further comprises
washing the analyte with a buffer solution applied at the second side under a mobilization field which moves the buffer solution from the second side to the first side.
14. The method of claim 13 , wherein the direction of the mobilization field is from a wash reservoir, which may be the same or different from the first buffer reservoir, to a wash waste reservoir, which may be the same or different from the first waste reservoir.
15. The method of claim 9 , which further comprises
adding a reagent to the analyte concentrated on the second side by concentrating the reagent on the second side with a mobilization field that crosses the size-exclusion polymeric element from the second side to the first side, wherein the mobilization field may be the same or different from the first electric field.
16. The method of claim 9 , which further comprises
loading the sample from the loading channel into the porous polymeric element by applying a mobilization field that does not cross the size-exclusion polymeric element.
17. The method of claim 16 , wherein the mobilization field is from the loading channel to the second waste reservoir.
18. The method of claim 9 , which further comprises
loading the sample from the loading channel into the porous polymeric element by applying a mobilization field that crosses the size-exclusion polymeric element.
19. The method of claim 9 , which further comprises
processing or assaying the analyte in the porous polymeric element by electrophoretic separation, chromatography, electrochromatography, immunochemistry, or a combination thereof.
20. The method of claim 19 , wherein the electrophoretic separation is capillary zone electrophoresis, capillary gel electrophoresis, native PAGE, SDS-PAGE, or a combination thereof.
21. A method for processing an analyte in a sample in a microfluidic device comprising a microfluidic channel having at least one in situ polymerized size-exclusion polymeric element having a first side and a second side, and at least one porous polymeric element adjacent to the second side, wherein the size-exclusion polymeric element has a length that is thinner than the length of the porous polymeric element, a loading channel in fluidic communication with a portion of the microfluidic channel containing the porous polymeric element, a sample reservoir in fluidic communication with a first waste reservoir, said sample reservoir is configured for containing a sample, wherein the sample reservoir and the first waste reservoir are configured to apply a first electric field from the sample reservoir through the loading channel and from the second side to the first side to the first waste reservoir, and a first buffer reservoir and a second waste reservoir configured to apply a second electric field from the first buffer reservoir and from the second side to the second waste reservoir through the length of the porous polymeric element, wherein the second electric field bypasses the size-exclusion polymeric element, wherein the method comprises
subjecting the sample to the microfluidic device to concentrate the analyte, filter the sample, exchange buffer, mix, react or bind the analyte with a reagent, or a combination thereof, and
moving the sample or the analyte through the porous polymeric element.