Nucleic acid sample preparation
The present invention includes methods, devices and systems for isolating a nucleic acid from a fluid comprising cells. In various aspects, the methods, devices and systems may allow for a rapid procedure that requires a minimal amount of material and/or results in high purity nucleic acid isolated from complex fluids such as blood or environmental samples.
1. A method for isolating a nanoparticulate from a sample, the method comprising:
a. applying the sample to a device, the device comprising an array of electrodes capable of establishing an AC electrokinetic field region;
b. concentrating cells and/or other particulate material in the sample in a first AC electrokinetic field region, wherein the first AC electrokinetic field region is a dielectrophoretic low field region and the conductivity is 300 mS/m or greater;
c. isolating the nanoparticulate in a second AC electrokinetic field region, wherein the second AC electrokinetic field is a dielectrophoretic high field region; and
d. flushing the concentrated cells and/or other particulate material from the first AC electrokinetic field region.
2. The method of claim 1 , wherein the AC electrokinetic field is produced using an alternating current having a voltage of 1 volt to 40 volts peak-peak, and/or a frequency of 5 Hz to 5,000,000 Hz and duty cycles from 5% to 50%.
3. The method of claim 1 , wherein the nanoparticulate is a nucleic acid, and the nucleic acid is deoxyribonucleic acid (DNA), ribonucleic acid (RNA) or a combination thereof.
4. The method of claim 1 , wherein the electrodes are selectively energized to provide the first dielectrophoretic field region and subsequently or continuously selectively energized to provide the second dielectrophoretic field region.
5. The method of claim 1 , wherein the array of electrodes is spin-coated with a hydrogel having a thickness between about 0.1 microns and 1 micron.
6. The method of claim 5 , wherein the hydrogel comprises two or more layers of a synthetic polymer.
7. The method of claim 5 , wherein the hydrogel has a viscosity between about 0.5 cP to about 5 cP prior to spin-coating.
8. The method of claim 5 , wherein the hydrogel has a conductivity between about 0.1 S/m to about 1.0 S/m.
9. The method of claim 1 , wherein the isolated nanoparticulate comprises less than 10% non-nucleic acid cellular material or cellular protein by mass.
10. The method of claim 1 , wherein the method is completed in less than 10 minutes.
11. The method of claim 1 , wherein the array of electrodes comprises a wavy line configuration, wherein the configuration comprises a repeating unit comprising the shape of a pair of dots connected by linker, wherein the linker tapers inward toward the midpoint between the pair of dots, wherein the diameters of the dots are the widest points along the length of the repeating unit, wherein the edge to edge distance between a parallel set of repeating units is equidistant, or roughly equidistant.
12. The method of claim 1 , wherein the array of electrodes comprises a passivation layer with a relative electrical permittivity from about 2.0 to about 4.0.
13. The method of claim 1 , wherein the sample is a bodily fluid, an environmental sample, food or beverage, growth medium or water.
14. A method for isolating a nanoparticulate from a sample, the method comprising:
a. applying the sample to a device, the device comprising an array of electrodes capable of establishing an AC electrokinetic field region;
b. concentrating cells and/or other particulate material in the sample in a first AC electrokinetic field region, wherein the first AC eletrokinetic field region is a dielectrophoretic low field region and the conductivity is 300 mS/m or greater;
c. isolating the nanoparticulate in a second AC electrokinetic field region, wherein the second AC electrokinetic field is a dielectrophoretic high field region;
d. flushing the concentrated cells and/or other particulate material from the first AC electrokinetic field region;
e. degrading residual proteins and/or material; and
f. flushing the degraded residual proteins and/or material from the isolated nanoparticulates.
15. The method of claim 14 , wherein the residual proteins and/or material are degraded by chemical and/or enzymatic degradation agents.
16. The method of claim 14 , wherein the AC electrokinetic field is produced using an alternating current having a voltage of 1 volt to 40 volts peak-peak, and/or a frequency of 5 Hz to 5,000,000 Hz and duty cycles from 5% to 50%.
17. The method of claim 14 , wherein the electrodes are selectively energized to provide the first dielectrophoretic field region and subsequently or continuously selectively energized to provide the second dielectrophoretic field region.
18. The method of claim 14 , wherein the array of electrodes is spin-coated with a hydrogel having a thickness between about 0.1 microns and 1 micron.
19. The method of claim 14 , wherein the array of electrodes comprises a wavy line configuration, wherein the configuration comprises a repeating unit comprising the shape of a pair of dots connected by linker, wherein the linker tapers inward toward the midpoint between the pair of dots, wherein the diameters of the dots are the widest points along the length of the repeating unit, wherein the edge to edge distance between a parallel set of repeating units is equidistant, or roughly equidistant.
20. The method of claim 14 , wherein the array of electrodes comprises a passivation layer with a relative electrical permittivity from about 2.0 to about 4.0.
21. The method of claim 14 , wherein the sample is a bodily fluid, an environmental sample, food or beverage, growth medium or water.