Method and apparatus for purifying nucleic acids and performing polymerase chain reaction assays using an immiscible fluid
Embodiments of the present invention are directed toward devices, systems, and methods for purifying nucleic acids to conduct polymerase chain reaction (PCR) assays. In one example, a method includes generating complexes of silica beads and nucleic acids in a lysis buffer, transporting the complexes through an immiscible fluid to remove interfering compounds from the complexes, further transporting the complexes into a density medium containing components required for PCR where the nucleic acids disassociate from the silica beads, and thermocycling the contents of the density medium to achieve PCR. Signal may be detected from labeling agents in the components required for PCR.
1. An apparatus for conducting an assay, the apparatus comprising:
a. a first fluidic feature which contains a fluid sample, wherein the fluid sample includes a plurality of complexes, individual ones of the complexes comprising a plurality of nucleic acids and a bead;
b. a second fluidic feature adjacent to said first fluidic feature, said second fluidic feature containing an immiscible fluid having a density greater than that of said fluid sample but lower than said complexes; and
c. a third fluidic feature adjacent to said second fluidic feature, which contains a density medium, wherein the density medium contains the components necessary for polymerase chain reaction (PCR) and said density medium has a density greater than said immiscible fluid but lower than said complexes.
2. The apparatus of claim 1 , further comprising a first control layer between the first and second fluidic features and a second control layer between the second and third fluidic features, said first and second control layers configured to be broken to create fluid communication between the first and second fluidic features and the second and third fluidic features.
3. The apparatus of claim 1 , wherein said fluidic features are portions of the same channel.
4. The apparatus of claim 1 , wherein said fluidic features are separate chambers in fluid communication.
5. The apparatus of claim 1 , wherein said fluidic features are contained on a microfluidic disk.
6. The apparatus of claim 1 , wherein the third fluidic feature comprises a first segment holding freeze dried PCR components and a second segment containing the density media, said first and second segments being separated by a third control layer, said third control layer configured to be broken to create fluid communication between the first and second segments of said third fluidic feature.
7. The apparatus of claim 1 , wherein the second fluidic feature's bottom surface is lower than said first fluidic feature's bottom surface and the third fluidic feature's bottom surface is lower than said second fluidic feature's bottom surface.
8. A system for conducting an assay, the system comprising:
a. an apparatus comprising:
i. a first fluidic feature which contains a fluid sample, wherein the fluid sample includes a plurality of complexes, individual ones of the complexes comprising a plurality of nucleic acids and a bead;
ii. a second fluidic feature adjacent to said first fluidic feature, said second fluidic feature containing an immiscible fluid having a density greater than that of said fluid sample but lower than said complexes;
iii. a third fluidic feature adjacent to said second fluidic feature, which contains a density medium, wherein the density medium contains the components necessary for polymerase chain reaction (PCR) and said density medium has a density greater than said immiscible fluid but lower than said complexes;
b. a motor coupled to the apparatus, the motor configured to receive a motor control signal and spin the apparatus responsive to the motor control signal;
c. a heating element in contact with the lower surface of the apparatus, the heating element configured to receive a heater control signal and heat the apparatus responsive to the heater control signal;
d. a detection module positioned to detect a signal from label agents included in the components necessary for PCR, wherein the detection module is configured to generate an electronic detection signal based, at least in part, on the signal from the label agents; and
e. a processing device coupled to the motor, the heating element, and the detection module, wherein the processing device is configured to allow an incubation period, generate the motor control signal and provide the motor control signal to the motor, generate the heater control signal and provide the heater control signal to the heating element, and wherein the processing device is further configured to receive the electronic detection signal from the detection module.
9. The system of claim 8 , wherein the apparatus further comprises a first control layer between the first and second fluidic features and a second control layer between the second and third fluidic features, said first and second control layers configured to be broken to create fluid communication between the first and second fluidic features and the second and third fluidic features.
10. The system of claim 9 , wherein a processing device is further configured to break the control layers by sending a heater control signal to the heating element.
11. The system of claim 10 , wherein the third fluidic feature further comprises a first segment holding freeze dried PCR components and a second segment containing the density media, said first and second segments being separated by a third control layer, said third control layer configured to be broken to create fluid communication between the first and second segments of said third fluidic feature.
12. The system of claim 8 , wherein said fluidic features are contained on a microfluidic disk.
13. The apparatus of claim 1 , wherein said bead has a density greater than that of said immiscible fluid.
14. The apparatus of claim 1 , wherein said complexes are configured to be transported through the immiscible fluid by way of centrifugal force.
15. The apparatus of claim 1 , wherein said complexes are configured to be transported into the density medium by way of centrifugal force.
16. The apparatus of claim 1 , wherein said complexes are configured to be transported through said first fluidic feature to said second fluidic feature by way of centrifugal force.
17. The apparatus of claim 16 , wherein said complexes are configured to be transported through said second fluidic feature to said third fluidic feature by way of centrifugal force.
18. The apparatus of claim 1 , further configured to effect transport of said complexes through the immiscible fluid and into a density medium by way of centrifugal force arising from spinning the apparatus.
19. The system of claim 8 , wherein said complexes are configured to be transported through the immiscible fluid by way of centrifugal force.
20. The system of claim 8 , wherein said complexes are configured to be transported into the density medium by way of centrifugal force.