Methods and apparatuses for processing blood and other biological samples
Systems and methods for the high yield extraction and recovery of higher quality target molecule(s) (e.g., DNA, RNA, protein, lipids, metabolites) from blood spots. High quality DNA recovered from dried blood spots can be an input source for high throughput analytical methods, such as for polymerase chain reaction (e.g., qPCR) and/or next generation sequencing (NGS). In various embodiments, at least 20.0 nanograms of nucleic acid (e.g., DNA, RNA), or at least 8.0 milligrams of protein, may be extracted and recovered per an amount of dried blood corresponding to approximately 5 microliters of fresh blood. In some embodiments, a majority (e.g., greater than 50%, greater than 60%, up to 95-100%) of the nucleic acid that is extracted and recovered from the blood spot via focused acoustics may be of a quality suitable for amplification via PCR or NGS.
1. A method of processing a dried blood spot, comprising:
placing a sample in a vessel, the sample including blood dried on to a substrate and a liquid; and
transmitting focused acoustic energy through a wall of the vessel such that the blood and the substrate are exposed to acoustic energy having a frequency of between about 100 kilohertz and about 100 megahertz at a focal zone having a size dimension of less than about 2 centimeters, resulting in extraction of biomolecules from the blood, wherein at least 60% of nucleic acid of the extracted biomolecules is capable of amplification via polymerase chain reaction.
2. A method of processing a dried blood spot, comprising:
placing a sample in a vessel, the sample including dried blood on a substrate and a liquid;
transmitting focused acoustic energy through a wall of the vessel such that the blood and the substrate are exposed to acoustic energy having a frequency of between about 100 kilohertz and about 100 megahertz at a focal zone having a size dimension of less than about 2 centimeters, resulting in extraction of biomolecules from the blood; and
recovering from the extracted biomolecules at least 20.0 nanograms of nucleic acid per an amount of dried blood corresponding to 5 microliters of fresh blood, or recovering from the extracted biomolecules at least 8.0 milligrams of protein per an amount of dried blood corresponding to 5 microliters of fresh blood.
3. The method of claim 2 , wherein at least 60% of nucleic acid of the extracted biomolecules is capable of amplification via polymerase chain reaction.
4. The method of claim 2 , wherein a greater percentage of nucleic acid extracted via the focused acoustic energy is capable of amplification via polymerase chain reaction than nucleic acid extracted without the focused acoustic energy.
5. The method of claim 2 , wherein at least 20.0 nanograms of nucleic acid of the extracted biomolecules per an amount of dried blood corresponding to 5 microliters of fresh blood is capable of amplification via polymerase chain reaction.
6. The method of claim 2 , wherein recovering from the extracted biomolecules includes recovering between 20.0 nanograms and 160.0 nanograms of nucleic acid per an amount of dried blood corresponding to 5 microliters of fresh blood.
7. The method of claim 2 , wherein recovering from the extracted biomolecules includes recovering between 8.0 milligrams and 20.0 milligrams of protein per an amount of dried blood corresponding to 5 microliters of fresh blood.
8. The method of claim 2 , wherein the sample includes an amount of dried blood corresponding to less than 15 microliters of fresh blood.
9. The method of claim 2 , wherein transmitting focused acoustic energy through the wall of the vessel includes exposing the blood and the substrate to focused acoustic energy for less than 10 minutes.
10. The method of claim 2 , wherein the liquid includes a buffer.
11. The method of claim 2 , wherein exposure of the extracted biomolecules to focused acoustic energy causes fragmenting of nucleic acid from the extracted biomolecules to an average size of less than 1000 bp.
12. The method of claim 2 , wherein the transmitting focused acoustic energy includes operating an acoustic transducer at a peak intensity power of 100 W to 300 W.
13. The method of claim 2 , wherein the transmitting focused acoustic energy includes operating an acoustic transducer at a duty factor of 10% to 90%.
14. The method of claim 2 , wherein the transmitting focused acoustic energy includes operating an acoustic transducer at a cycles per burst of 100 to 300.
15. The method of claim 2 , wherein the transmitting focused acoustic energy includes operating an acoustic transducer at a peak intensity power of between 100 W and 300 W, a duty factor of between 10% and 90% and a cycles per burst of between 100 and 300.
16. The method of claim 2 , further comprising incubating the sample within the vessel at a temperature of between 20 degrees C. and 60 degrees C.
17. The method of claim 16 , further comprising incubating the sample within the vessel at a temperature of between 50 degrees C. and 60 degrees C.
18. The method of claim 16 , further comprising incubating the sample within the vessel at a temperature of between 20 degrees C. and 40 degrees C.
19. A method of processing a dried blood spot, comprising:
placing a sample in a vessel, the sample including blood dried on to a substrate and a liquid; and
transmitting focused acoustic energy through a wall of the vessel such that the blood and the substrate are exposed to acoustic energy having a frequency of between about 100 kilohertz and about 100 megahertz at a focal zone having a size dimension of less than about 2 centimeters, resulting in extraction of biomolecules from the blood, wherein a greater percentage of nucleic acid extracted from the sample via the focused acoustic energy is capable of amplification via polymerase chain reaction than nucleic acid extracted from an identical sample using extraction protocol from a QIAamp DNA Mini Kit without the focused acoustic energy.
20. The method of claim 1 , wherein at least 20.0 nanograms of nucleic acid of the extracted biomolecules per an amount of dried blood corresponding to 5 microliters of fresh blood is capable of amplification via polymerase chain reaction.
21. The method of claim 1 , wherein transmitting focused acoustic energy through the wall of the vessel includes exposing the blood and the substrate to focused acoustic energy for less than 10 minutes.
22. The method of claim 1 , wherein transmitting focused acoustic energy through the wall of the vessel causes fragmenting of nucleic acid from the extracted biomolecules to an average size of less than 1000 bp.
23. The method of claim 1 , wherein transmitting focused acoustic energy through the wall of the vessel includes operating an acoustic transducer at a peak intensity power of between 100 W and 300 W, a duty factor of between 10% and 90% and a cycles per burst of between 100 and 300.