Sample preparation device and methods of use
A device for isolating DNA from a sample containing cells, including a cartridge having an entrance port and an exit port, a membrane disposed between the entrance port and the exit port, and a plurality of channels between the membrane and the exit port. Additionally, systems and methods for isolating DNA from a sample containing cells and also systems and methods for amplifying and isolating single-stranded DNA from a sample containing DNA.
1. A device for isolating DNA from a sample containing cells, comprising
a cartridge having an entrance port and an exit port;
a membrane disposed between the entrance port and the exit port;
a plurality of channels between the membrane and the exit port; and
a plurality of particulate material configured to mechanically lyse cells, wherein the plurality of particulate material is located between the membrane and the entrance port.
2. The device of claim 1 , wherein the channels are adjacent to the membrane.
3. The device of claim 1 , wherein each of the channels has a channel entrance, a channel exit, and a channel wall, wherein the channel entrance is adjacent to the membrane.
4. The device of claim 3 , wherein the channel entrance has a length and width, wherein the length is about 0.1 mm to about 100 mm and the width is about 0.1 mm to about 10 mm.
5. The device of claim 1 , having from about 2 to about 1000 channels.
6. The device of claim 1 , wherein the channels are arranged in a matrix.
7. The device of claim 6 , wherein the channels are adjacent to each other.
8. The device of claim 6 , wherein the channels are spaced apart from each other.
9. The device of claim 8 , further comprising a solid barrier between each channel entrance.
10. The device of claim 1 , wherein the membrane is configured to be permeable to DNA but substantially impermeable to cells.
11. The device of claim 1 , wherein the membrane is configured to be permeable to DNA but substantially impermeable to non-DNA cellular material.
12. The device of claim 1 , wherein the membrane comprises a cellulose material.
13. The device of claim 1 , wherein the membrane has pores.
14. The device of claim 13 , wherein the pores are from about 10 nm to 1 micron in diameter.
15. The device of claim 1 , wherein the particulate material comprises glass or stainless steel.
16. The device of claim 1 , wherein the particulate material comprises a plurality of microbeads.
17. The device of claim 16 , wherein the microbeads have a diameter of from about 1 micron to about 1000 microns.
18. The device of claim 1 , wherein the entrance port and exit port comprise standard luer lock fittings.
19. The device of claim 1 , wherein the cartridge includes a bottom piece that supports the membrane and the plurality of channels are formed from cavities in the bottom piece of the cartridge.
20. A system for isolating DNA from a sample containing cells comprising the device of claim 1 , and a vacuum to pull fluid into the cartridge.
21. The system of claim 20 , further comprising a cellular homogenizer to shake the cartridge.
22. The system of claim 21 , wherein the cellular homogenizer is capable of shaking the cartridge at a rate of up to 5000 rpm.
23. The system of claim 21 , further comprising a thermal cycler configured to subject the DNA to thermal cycling for a polymerase chain reaction (PCR).
24. The system of claim 23 , further comprising a magnetic substrate wherein the magnetic substrate is a coating with a binding affinity for double-stranded DNA.
25. The system of claim 24 , further comprising a magnet configured to apply a magnetic field to the magnetic substrate.
26. The system of claim 24 , wherein the magnetic substrate is a plurality of magnetic beads.
27. A method of processing DNA from a sample, comprising
providing a sample containing cells having DNA;
transferring the sample into the device of claim 1 ;
lysing the cells to release the DNA and non-DNA cellular material;
passing the DNA through the membrane; and
passing the DNA through the channels;
wherein the DNA is isolated from the sample.
28. The method of claim 27 , wherein the sample comprises blood, exudate, water, or food.
29. The method of claim 27 , wherein the transferring comprises using a vacuum to transfer the sample into the cartridge.
30. The method of claim 27 , wherein the mechanical lysing substantially destroys the non-DNA cellular material.
31. The method of claim 27 , wherein greater than 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the non-DNA cellular material in the sample does not pass through the membrane.
32. The method of claim 27 , wherein greater than 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the DNA in the sample is passed through the membrane and the channels.
33. The method of claim 27 , further comprising transferring the isolated DNA to a DNA amplification chamber and amplifying the isolated DNA by a polymerase chain reaction (PCR), wherein the PCR is asymmetric, to produce double-stranded DNA and single-stranded DNA.
34. The method of claim 33 , wherein a portion of the single-stranded DNA is within a specified range of lengths.
35. The method of claim 33 , further comprising binding the double-stranded DNA to a magnetic substrate and applying a magnetic field to the substrate, wherein the magnetic field attracts the substrate bound to the double-stranded DNA, and isolating the single-stranded DNA.