High resolution focusing and separation of proteins in nanofluidic channels
Exemplary embodiments provide systems and methods for concentrating, focusing and/or separating proteins using nanofluidic channels and/or their arrays. In embodiments, low-abundance proteins can be focused and separated with high resolution using separation techniques including isoelectric focusing (IEF), and/or dynamic field gradient focusing (DFGF) in combination with nanofluidic channels and/or multi-gate nanofluidic field-effect-transistors (FETs).
1. A method for focusing proteins comprising:
providing a plurality of nanofluidic channels, wherein one of a width and a depth of each nanofluidic channel is about 1000 nm or less;
equilibrating a buffer solution with walls of the plurality of nanofluidic channels suitable for the proteins to be focused by determining an equilibrium time by IR absorbance spectra;
introducing a protein mixture solution that contains a plurality of proteins into the nanofluidic channels; and
applying an electric potential to a length of the protein mixture solution along the nanofluidic channels to generate a longitudinal electric field.
2. The method of claim 1 , further comprising forming at least one focused protein band by the longitudinal electric field, wherein the at least one focused protein band has a width of about 100 micrometers or less.
3. The method of claim 2 , wherein the at least one focused protein band is statically focused for a period of time ranging from about 5 minutes to about 30 minutes.
4. The method of claim 2 further comprising advancing a location of the at least one focused protein band further along the nanofluidic channels by increasing the applied electric potential.
5. The method of claim 1 , wherein the protein mixture solution has a protein concentration of about 1 millimolar or less.
6. The method of claim 1 further comprising separating the plurality of proteins in the protein mixture solution using one or more of isoelectric focusing (IEF), dynamic field gradient focusing (DFGF) and a combination thereof.
7. The method of claim 1 further comprising applying a low electric potential of about 5 V or less for isoelectric focusing (IEF) the plurality of proteins in the protein mixture solution.
8. The method of claim 1 further comprising:
filling the buffer solution into the plurality of nanofluidic channels, and
equilibrating the buffer solution with walls of the plurality of nanofluidic channels.
9. The method of claim 1 further comprising increasing the applied electric potential to reduce a band formation time from the application of the electric potential.
10. The method of claim 1 further comprising determining and applying a high electric potential to generate the longitudinal electric field when the plurality of proteins in the protein mixture solution have a high molecular weight.
11. The method of claim 1 further comprising changing the electric potential applied to the protein mixture solution to change a flowing direction of the plurality of proteins along the nanofluidic channels.
12. The method of claim 1 further comprising separating a plurality of proteins with sizes in the same range.
13. The method of claim 1 further comprising configuring a multi-gate nanofluidic field-effect-transistor (FET) having a plurality of gates spaced along the nanofluidic channels.
14. The method of claim 13 further comprising applying a second electric potential through the plurality of gates of the multi-gate nanofluidic FET to dynamically control at least one of a pH gradient, an electric field gradient and electrokinetic transport of the plurality of proteins in the nanofluidic channels.